Novel substituted succinic acid metallo-beta-lactamase inhibitors and their use in treating bacterial infections

ABSTRACT

This invention relates to novel substituted succinic acid metallo-β-lactamase inhibitors which are useful potentiators of β-lactam antibiotics. Accordingly, the present invention provides a method of treating bacterial infections in animals or humans which comprises administering, together with a b-lactam antibiotic, a therapeutically effective amount of a compound of formula I:  
                 
 
     including pharmaceutically acceptable salts, prodrugs, anhydrides, and solvates thereof.

BACKGROUND OF THE INVENTION

[0001] The present invention relates to compounds which havemetallo-β-lactamase inhibitory characteristics. The invention alsorelates to methods of preparing, pharmaceutical compositions and uses ofthe compounds.

[0002] Metallo-β-lactamases are bacterial enzymes which conferresistance to virtually all clinically relevant β-lactam antibiotics,including carbapenems and jeopardize the future use of all such agents.The increased treatment of infections with carbapenems and otherβ-lactam antibiotics may lead to the proliferation of clinical bacterialstrains which are able to produce metallo-β-lactamases and thus resistthe effects of β-lactam antibiotics. In fact, metallo-β-lactamases havenow been identified in a number of pathogenic bacterial speciesincluding Bacillus cereus, Bacteroides fragilis, Aeromonas hydrophila,Klebsiella pneumoniae, Pseudomonas aeruginosa, Serratia marcescens,Stenotrophomonas maltophilia, Shigella flexneri, Legionella gormanii,Chryseobacterium meningosepticum, Chryseobacterium indologenes,Acinetobacter baumannii, Citrobacter freundii, and Aeromonas veronii.

[0003] Accordingly, there is an increasing need for agents which whencombined with a β-lactam antibiotic, e.g. imipenem, will restore theeffectiveness of the β-lactam antibiotics and which are at the same timerelatively free from undesirable side effects.

[0004] WO 98/17639, 97/30027, 98/40056,98/39311 and 97/10225 teachcertain beta-thiopropionyl-amino acid derivatives and their use asinhibitory agents against metallo-β-lactamases. Goto et. al., Biol.Pharm. Bull. 20, 1136 (1997), Payne et. al., FEMS Microbiology Letters157, 171 (1997), Payne et al., Antimicrob. Agents Chemother. 41, 135(1997), Page et. al., Chem. Commun. 1609 (1998) and Page et al.,Biochem. J. 331, 703 (1998) also disclose certain thiols and thioestersas metallo-β-lactamase inhibitors. Additionally, Toney et al., Chemistryand Biology 5, 185 (1998), Fastrez et al., Tetrahedron Lett. 36, 9313(1995), Schofield et al., Tetrahedron 53, 7275 (1997), Schofield et.al., Bioorg. & Med. Chem. Lett. 6, 2455 (1996) and WO 97/19681 discloseother metallo-β-lactamase inhibitors. However, the above notedreferences do not teach the compounds of the instant invention.

[0005] Other references which disclosed the general state of the art areBush et al., Antimicrob. Agents Chemother. 41, 223 (1997); Livermore, D.M. J. Antimicrob. Chemother. 1998, 41 (Suppl. D), 25; Bush, K. Clin.Infect. Dis. 1998, 27 (Suppl 1), S48; Livermore, D. M. J. Antimicrob.Chemother. 1997, 39, 673 and Payne, D. J. J. Med. Microbiol. 1993, 39,93.

SUMMARY OF THE INVENTION

[0006] This invention relates to novel substituted succinic acidmetallo-β-lactamase inhibitors, which are useful potentiators ofβ-lactam antibiotics. Accordingly, the present invention provides amethod of treating bacterial infections in animals or humans whichcomprises administering, together with a β-lactam antibiotic, atherapeutically effective amount of a compound of formula I:

[0007] including pharmaceutically acceptable salts, prodrugs,anhydrides, and solvates thereof, wherein:

[0008] M¹ and M² are independently selected from:

[0009] (a) Hydrogen,

[0010] (b) Pharmaceutically acceptable cation,

[0011] (c) Pharmaceutically acceptable esterifying group; and

[0012] (d) A negative charge;

[0013] R¹ and R² are independently selected from the following:

[0014] (a) Hydrogen, provided that R¹ and R² are not hydrogen at thesame time;

[0015] (b) a C₁ to C₁₆ straight, branched or unsaturated alkyl groupsubstituted with 0 to 2 Rq groups and substituted with 0 to 3 R_(x)groups and optionally interrupted by one of the following O, S, SO₂,—C(O)—,

[0016] (c) —C(O)—NR^(a)—, —CO₂—;

[0017] (c) a group of the formula:

[0018]  wherein

[0019] —A— represents a single bond, C₁ to C₈ straight, branched orunsaturated alkyl group optionally substituted with 1 to 2 R_(x) groupsand optionally interrupted by one of the following O, S, SO₂, —C(O)—,—C(O)—NR^(a)—, —CO₂—;

[0020] represents:

[0021] (1) a C₆ to C₁₄ aryl group;

[0022] (2) a C₃ to C₁₀ alicyclic group;

[0023] (3) a C₃ to C₁₄ heteroaryl group, which contains 1 to 3heteroatoms, 0 to 3 of which heteroatoms are nitrogen and 0 to 1 ofwhich are oxygen or sulfur;

[0024] (4) a C₃ to C₁₀ heterocyclic group, which contains 1 to 2heteroatoms, 0 to 1 of which heteroatoms are nitrogen, and 0 to 2 ofwhich are oxygen or sulfur; or

[0025] (d) a group of the formula:

[0026]  wherein:

[0027] —A— is as defined above;

[0028] A′ is a single bond, O, S, or a C₁ to C₆ straight, branched orunsaturated alkyl group optionally substituted with 1-2 R_(x) groups andoptionally interrupted by one of the following groups O, S, SO₂, —C(O)—,—C(O)—NR^(a)—, —CO₂—;

[0029] are independently selected from:

[0030] (1) a C₆ to C₁₀ aryl group;

[0031] (2) a C₃ to C₈ alicyclic group;

[0032] (3) a C₂ to C₉ heteroaryl group, which contains 1 to 3heteroatoms, 0 to 3 of which heteroatoms are nitrogen and 0 to 1 ofwhich are oxygen or sulfur,

[0033] (4) a C₃ to C₈ heterocyclic group, which contains 1 to 2heteroatoms, 0 to 1 of which heteroatoms are nitrogen, and 0 to 2 ofwhich are oxygen or sulfur;

[0034] provided that at least one Rq group is present in R¹ or R² andthat when more than one Rq is present the total number of cationicnitrogen atoms does not exceed 8; the total number of cationic nitrogenatoms can be charged balanced by M¹ and/or M² or by M¹ and/or M² incombination with an appropriate number of Y⁻;

[0035] wherein:

[0036] Rq is —E—Q⁺Y⁻;

[0037] Y⁻ is a pharmaceutically acceptable anionic group;

[0038] E is —(CH₂)_(m)—X—(CH₂)_(n)—;

[0039] m is 0 to 6;

[0040] n is 0 to 6 (but when E is attached to an aromatic ring n is1-6);

[0041] X is a bond, O, S, SO₂, —C(O)—, —C(O)—N(R^(a))—, —C(O)O—, —CH=CH—or—C—C—, provided that when X is O, S, —C(O)—N(R^(a))— or —C(O)O—, thenn is 2 to 6

[0042] and Q⁺, attached to the (CH₂)n terminus of E is:

[0043] (1) a cationic group selected from the following:

[0044] wherein:

[0045] R^(u) and R^(v) are independently hydrogen or C₁₋₆ alkyloptionally substituted with 1 to 2 R^(y);

[0046] R^(w) is hydrogen or C₁₋₆ alkyl optionally substituted with 1 to2 R_(x);

[0047] R^(u) and R^(v) when bonded to the same nitrogen atom maytogether be a C₃₋₆ alkyl radical, which when taken together with theintervening atoms form a ring;

[0048] Two R^(u) groups on separate nitrogen atoms may together comprisea C₂₋₅ alkyl radical, which when taken together with the interveningatoms form a ring;

[0049] R^(u), R^(v) and R^(w) when bonded to the same nitrogen atom maytogether form a C₆₋₁₀ tertiary alkyl radical, which with N⁺ forms abicyclic ring;

[0050] (2) A dicationic group:

[0051] wherein:

[0052] E¹ is —(CH₂)_(p)—Z—(CH₂)_(r)—;

[0053] p and r are independently 1 to 4;

[0054] Z is a bond, O, S, SO₂, —C(O)—, —C(O)O—**, —CH═CH—, —C═C—, or

[0055] Provided that when Z is O or S, p is 2 to 4 and r is 2 to 4 andwhen Z is

[0056] or —C(O)O—**, r is 2 to 4;

[0057] wherein ** denotes the atom which is bonded to the —(CH₂)_(r)—moiety of E¹ above;

[0058] Q¹ is selected from the following:

[0059]  Q² is selected from the following:

[0060] R^(u), R^(v) and R^(w) are independently selected and defined asabove,

[0061] And in addition, in the case where two R^(u) groups on separatenitrogen atoms are joined to form a ring as defined above, two R^(v)groups on the same two separate nitrogen atoms may also comprise a C₁₋₅alkyl radical to form together with the intervening atoms a bicyclicring; an example of such is:

[0062] (3) A tricationic group selected from the following:

[0063]  wherein:

[0064] Each E¹ is as defined above, but selected independently;

[0065] Each Q¹ is as defined above, but selected independently;

[0066] Each Q² is as defined above, but selected independently;

[0067] R^(u), R^(v) and R^(w) are defined as in the definition of Q⁺item (2) above and selected independently; or

[0068] (4) A tetracationic group selected from the following:

[0069]  wherein:

[0070] Each E¹ is as defined above, but selected independently;

[0071] Each Q¹ is as defined above, but selected independently;

[0072] Each Q² is as defined above, but selected independently;

[0073] R^(u), R^(v) and R^(w) are defined as in the definition of Q⁺item (2) above and selected independently;

[0074] where each R_(x) is independently selected from the groupconsisting of:

[0075] (a) F, Cl, Br, I,

[0076] (b) CF₃,

[0077] (c) OR^(b),

[0078] (d) CN,

[0079] (e) —C(O)—R^(c),

[0080] (f) —S(O₂)—R^(f),

[0081] (g) —C(O)—OR^(a)

[0082] (h) —O—C(O)—R^(c),

[0083] (i) —S—R^(b),

[0084] (j) —N(R^(a))—C(O)—R^(c),

[0085] (q) —N(R^(a))—C(O)—OR^(f),

[0086] (r) —S(O)—R^(f),

[0087] (s) —N(R^(a))—S(O₂)—R^(f),

[0088] (t) NO₂, and

[0089] (u) C₁ to C₈ straight, branched or unsaturated alkyl optionallysubstituted with one of the substituents (a) through (t) above;

[0090] (v) —CH₂-aryl wherein the aryl is optionally substituted with oneof the substituents (a) through (t) above;

[0091] or two adjacent R_(x) groups on an aromatic ring may consist ofthe following divalent moiety, —O—CH₂—O—;

[0092] wherein:

[0093] R^(a) is H, C₁ to C₆ alkyl optionally substituted with R^(y);

[0094] R^(b) is H, C₁ to C₆ alkyl optionally substituted with R^(y),CH₂-aryl, or aryl, said aryls optionally substituted with 1-2 R^(y)groups;

[0095] R^(c) is H, C₁ to C₆ alkyl optionally substituted with R^(y),CF₃, or aryl, said aryl optionally substituted with 1-2 R^(y) groups;

[0096] R^(d) and R^(e) are independently hydrogen, C₁ to C₄ alkyloptionally substituted with R^(y), or R^(d) and R^(e) taken together mayrepresent a 3 to 5-membered alkyl radical to form a ring, or R^(d) andR^(e) taken together may represent a 2 to 4-membered alkyl radicalinterrupted by O, S, SO or SO₂ to form a ring;

[0097] R^(f) is C₁ to C₆ alkyl optionally substituted with R^(y), oraryl, said aryl optionally substituted with 1-2 R^(y) groups; and

[0098] R^(y) is —OH, —OCH₃, OCONH₂, OCOCH₃, CHO, COCH₃, CO₂CH₃, CONH₂,CN, SOCH₃, SO₂CH₃, SO₂NH₂, F, Cl, Br, I or CF₃.

[0099] The invention is intended to include all of the isomeric forms ofthe compounds of formula I, including racemic, enantiomeric anddiastereomeric forms.

DETAILED DESCRIPTION OF THE INVENTION

[0100] The invention is described herein in detail using the termsdefined below unless otherwise specified.

[0101] The term “alkyl” refers to a monovalent alkane (hydrocarbon)derived radical containing from 1 to 16 carbon atoms unless otherwisedefined. It may be straight or branched. Preferred alkyl groups includemethyl, ethyl, propyl, isopropyl, butyl and hexyl. When substituted,alkyl groups may be substituted with up to 3 substituent groups selectedfrom R_(x), as defined, and up to 2 substituent groups selected from Rq,as defined, at any available point of attachment. When the alkyl groupis said to be substituted with an alkyl group, this is usedinterchangeably with “branched alkyl group”. When the alkyl chain isinterrupted by a group, eg. O, this may occur between any two saturatedcarbons of the alkyl chain.

[0102] The term unsaturated alkyl refers to “alkenyl” or “atkynyl”. Theterm “alkenyl” refers to an unsaturated alkyl such as a hydrocarbonradical, straight or branched containing from 2 to 16 carbon atoms andat least one carbon to carbon double bond. Preferred alkenyl groupsinclude propenyl, hexenyl and butenyl. The term “alkynyl” refers to anunsaturated alkyl such as a hydrocarbon radical straight or branched,containing from 2 to 16 carbon atoms and at least one carbon to carbontriple bond. Preferred alkynyl groups include propynyl, hexynyl andbutynyl.

[0103] The term “alicyclic” refers to non-aromatic monocyclic orbicyclic C₃-C₁₀ hydrocarbons, including unsaturated, which can besubstituted with 0-3 groups of R_(x). Examples of said groups includecycloalkyls such as cyclohexyl, cyclopentyl, bicyclo[2.2.1]heptyl,bicyclo[2.2.1]hepta-2,5-dienyl, bicyclo[2.2.2]octyl,bicyclo[2.2.2]octa-2,5-dienyl.

[0104] The term “alkylidene” refers to an alkyl group which is attachedthrough two bonds on the same carbon atom of the alkyl group to a singleattachment atom Examples of said groups include methylene, ethylidene,isopropylidene and the like.

[0105] Examples of when R^(d) and R^(e) are taken together along withthe adjacent nitrogen atom to represent a 3 to 5 membered alkyl radicalforming a ring or a 2 to 4 membered alkyl radical interrupted by O, S,SO, SO₂, to form a ring are pyrrolidinyl, piperidinyl, morpholinyl andthe like.

[0106] The term “heterocyclic” refers to a monocyclic non-aromaticmoiety containing 3-8 ring atoms or a bicyclic non-aromatic moietycontaining 6-10 ring atoms, at least one of which ring atoms is aheteroatom selected from nitrogen, oxygen and sulfur and where oneadditional ring atom may be oxygen or sulfur. Examples of heterocyclicgroups are furanyl, pyranyl, morpholinyl, dioxanyl and quinuclidinyl:

[0107] Aryl refers to aromatic rings e.g., phenyl, substituted phenyland the like, as well as rings which are fused, e.g., naphthyl,phenanthrenyl fluorenonyl and the like. An aryl group thus contains atleast one ring having at least 6 atoms, with up to three such ringsbeing present, containing up to 14 atoms therein, with alternating(resonating) double bonds between adjacent carbon atoms. The preferredaryl groups are phenyl, naphthyl, and fluorenone. Aryl groups maylikewise be substituted as defined. Preferred substituted aryls includephenyl, fluorenonyl and naphthyl.

[0108] The term “heteroaryl” (Het) refers to a monocyclic aromatic grouphaving 5 or 6 ring atoms, a bicyclic aromatic group having 8 to 10atoms, or tricyclic having 12-14 ring atoms, containing at least oneheteroatom, O, S or N, in which a carbon or nitrogen atom is the pointof attachment, and in which one or two additional carbon atoms isoptionally replaced by a heteroatom selected from O or S, and in whichfrom 1 to 3 additional carbon atoms are optionally replaced by nitrogenheteroatoms, said heteroaryl group being optionally substituted asdescribed herein. Examples of this type are pyrrole, pyridine, oxazole,thiazole, dibenzofuran, dibenzothiophene, carbazole, phenanthrene,anthracene, dibenzothiophene sulfone, fluorenone, quinoline and oxazine.Additional nitrogen atoms may be present together with the firstnitrogen and oxygen or sulfur, giving, e.g., thiadiazole. Examplesinclude the following:

[0109] Heteroarylium refers to heteroaryl groups bearing a quaternarynitrogen atom and thus a positive charge. Non-limiting examples includethe following:

[0110] When a charge is shown on a particular nitrogen atom in a ringwhich contains one or more additional nitrogen atoms, it is understoodthat the charge may reside on a different nitrogen atom in the ring byvirtue of charge resonance that occurs.

[0111] Similar charge resonance may occur in amidinium and guanidimumgroups:

[0112] The term “heteroatom” means O, S or N, selected on an independentbasis.

[0113] Halogen and “halo” refer to bromine, chlorine, fluorine andiodine.

[0114] The term “pro-drug” refers to compounds with a removable groupattached to one or both of the carboxyl groups of compounds of formula I(e.g. biolabile esters). Groups which are useful in forming pro-drugsshould be apparent to the medicinal chemist from the teachings herein.Examples include pivaloyloxymethyl, acetoxymethyl, phthalidyl, indanyland methoxymethyl, and others described in detail in U.S. Pat. No.4,479,947. These are also referred to as “biolabile esters”.

[0115] The term “hydrate” is used in the conventional sense to includethe compounds of formula I in physical association with water.

[0116] When a group is termed “substituted”, unless otherwise indicated,this means that the group contains from 1 to 3 substituents thereon.

[0117] A bond terminated by a wavy line is used herein to signify thepoint of attachment of a substituent group. This usage is illustrated bythe following example:

[0118] The terms “quaternary nitrogen” and “cationic nitrogen” refer totetravalent, positively charged nitrogen atoms including, e.g., thepositively charged nitrogen in a tetraalkylammonium group (e. g.—N⁺R^(u)R^(v)R^(w)), heteroarylium, (e.g., N-methyl-imidazolium),amidinium, guanidinium, basic nitrogens which are protonated atphysiological pH, and the like. A “cationic group” is a moiety whichcontains at least one such quaternary nitrogen atom. Cationic groupsthus encompass positively charged nitrogen-containing groups, as well asbasic nitrogens which are protonated at physiologic pH. The termsdicationic, tricationic and tetracationic refer to groups which contain2, 3 or 4 positively charged nitrogen atoms, respectively.

[0119] When a functional group is termed “protected”, this means thatthe group is in modified form to preclude undesired side reactions atthe protected site. Suitable protecting groups for the compounds of thepresent invention will be recognized from the present application takinginto account the level of skill in the art, and with reference tostandard textbooks, such as Greene, T. W. et al. Protective Groups inOrganic Synthesis Wiley, New York (1991). Examples of suitableprotecting groups are contained throughout the specification.

[0120] In some of the compounds of the present invention suitableprotecting groups represents hydroxyl-protecting, amine-protecting orcarboxyl-protecting groups. Such conventional protecting groups consistof groups, which are used to protectively block the hydroxyl, amine orcarboxyl group during the synthesis procedures described herein. Theseconventional blocking groups are readily removable, i.e., they can beremoved, if desired, by procedures which will not cause cleavage orother disruption of the remaining portions of the molecule. Suchprocedures include chemical and enzymatic hydrolysis, treatment withchemical reducing or oxidizing agents under mild conditions, treatmentwith a transition metal catalyst and a nucleophile and catalytichydrogenation.

[0121] Examples of carboxyl protecting groups include allyl, benzhydryl,2-naphthylmethyl, benzyl, silyl such as t-butyldimethylsilyl (TBDMS),phenacyl, p-methoxybenzyl, o-nitrobenzyl, p-methoxyphenyl,p-nitrobenzyl, 4-pyridylmethyl and t-butyl.

[0122] Examples of suitable amine protecting groups include9-fluorenylmethoxycarbonyl, p-nitrobenzyloxycarbonyl, benzyloxycarbonyl,allyloxycarbonyl, t-butyloxycarbonyl, 2,2,2-trichloroethyloxycarbonyland the like.

[0123] Examples of suitable hydroxyl protecting groups includetriethylsilyl, t-butyldimethylsilyl, o-nitrobenzyloxycarbonyl,p-nitrobenzyloxycarbonyl, benzyloxycarbonyl, allyloxycarbonyl,t-butyloxycarbonyl, 2,2,2-trichloroethyloxycarbonyl and the like.

[0124] With respect to M¹ and/or M², this represents a carboxylichydrogen, a carboxylate anion (M represents a negative charge), apharmaceutically acceptable ester (M represents an ester forming group)or a pharmaceutically acceptable cation. When M¹ and/or M² is a negativecharge it can be used to provide the necessary charge balance in acompound with one or more positive charges. Likewise, when M¹ and/or M²is a negative charge it can be balanced by the appropriate number ofcounterions, e.g., an alkali metal cation such as sodium or potassium.Other pharmaceutically acceptable counterions may be calcium, magnesium,zinc, ammonium, or alkylammonium cations such as tetramethylammonium,tetrabutylammonium, choline, triethylhydroammonium, meglumine,triethanolhydroammonium, etc.

[0125] For the purposes of this invention, all compounds have at leastone Rq substituent containing at least one cationic nitrogen. Preferably2 to 8 cationic nitrogens, more preferably 2 to 4 cationic nitrogens andmost preferably 3 to 4 cationic nitrogens are present. The compounds arebalanced with one or more, as necessary, of a charge balancing group Y⁻.Alternatively, the compounds can be balanced using M¹ and/or M² as thecharge balancing group with or without the use of Y⁻. Examples of caseswhere a charge balancing group is required are quaternized substituentssuch as heteroarylium, —N⁺R^(u)R^(v)-E¹-Q¹, —N⁺R^(u)R^(v)R^(w), and thelike.

[0126] Additionally, all compounds having one or more anions are counterbalanced with one or more, as necessary, charge balancing cations.

[0127] The compounds of the present invention are useful per se and intheir pharmaceutically acceptable salt and ester forms are potentiatorsfor the treatment of bacterial infections in animal and human subjects.The term “pharmaceutically acceptable ester, salt or hydrate”, refers tothose salts, esters and hydrated forms of the compounds of the presentinvention which would be apparent to the pharmaceutical chemist. Forexample, those which are substantially non-toxic and which may favorablyaffect the pharmacokinetic properties of said compounds, such aspalatability, absorption, distribution, metabolism and excretion. Otherfactors, more practical in nature, which are also important in theselection, are cost of the raw materials, ease of crystallization,yield, stability, solubility, hygroscopicity and flowability of theresulting bulk drug. Conveniently, pharmaceutical compositions may beprepared from the active ingredients in combination withpharmaceutically acceptable carriers. Thus, the present invention isalso concerned with pharmaceutical compositions and methods of treatingbacterial infections utilizing as an active ingredient the novelcarbapenemase compounds.

[0128] The pharmaceutically acceptable salts referred to above alsoinclude acid addition salts. Thus, the Formula I compounds can be usedin the form of salts derived from inorganic or organic acids. Includedamong such salts are the following:

[0129] acetate, adipate, alginate, aspartate, benzoate,benzenesulfonate, bisulfate, butyrate, citrate, camphorate,camphorsulfonate, cyclopentanepropionate, digluconate, dodecylsulfate,ethanesulfonate, fumarate, glucoheptanoate, glycerophosphate,hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide,hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate,methanesulfonate, 2-naphthalenesulfonate, nicotinate, oxalate, pamoate,pectinate, persulfate, 3-phenylpropionate, picrate, pivalate,propionate, succinate, tartrate, thiocyanate, tosylate and undecanoate.

[0130] Acid addition salts of the compounds of formula I includecompounds that contain a protonated, basic moiety in R^(q). Compoundscontaining a basic moiety in R^(q) are capable of protonation in aqueousmedia near pH 7, so that the basic moiety can exist as an equilibriummixture of its neutral form and acid addition (protonated) form. Themore basic the group, the greater the degree of protonation near pH 7.All such compounds are included in the present invention.

[0131] The pharmaceutically acceptable cations which can form a saltwith one or both of the carboxyls (CO₂M¹ and CO₂M²) of the compounds offormula I are known to those skilled in the art. Examples include thosewhere M¹ and M² independently can be alkali metals such as sodium,potassium and the like, ammonium and the like.

[0132] The pharmaceutically acceptable esterifying groups are such aswould be readily apparent to a medicinal chemist, and include, forexample, those described in detail in U.S. Pat. No. 4,309,438. Includedwithin such pharmaceutically acceptable esters are those which arehydrolyzed under physiological conditions, such as pivaloyloxymethyl,acetoxymethyl, phthalidyl, indanyl and methoxymethyl, and othersdescribed in detail in U.S. Pat. No. 4,479,947. These are also referredto as “biolabile esters”.

[0133] Biolabile esters are biologically hydrolizable, and may besuitable for oral administration, due to good absorption through thestomach or intestinal mucosa, resistance to gastric acid degradation andother factors. Examples of biolabile esters include compounds in whichM¹ and/or M² represents an alkoxyalkyl, alkylcarbonyloxyalkyl,alkoxycarbonyloxyalkyl, cycloalkoxylalkyl, alkenyloxyalkyl,aryloxyalkyl, alkoxyaryl, alkylthioalkyl, cycloalkylthioalkyl,alkenylthioalkyl, arylthioalkyl or alkylthioaryl group. The following M¹and/or M² species are examples of biolabile ester forming moieties:acetoxymethyl, 1-acetoxyethyl, 1-acetoxypropyl, pivaloyloxymethyl,1-isopropyloxycarbonyloxyethyl, 1-cyclohexyloxycarbonyloxyethyl,phthalidyl and (2-oxo-5-methyl-1,3-dioxolen-4-yl)methyl.

[0134] Y⁻ can be present or absent as necessary to maintain theappropriate charge balance. When present, these representpharmaceutically acceptable counterions. Most anions derived frominorganic or organic acids are suitable. Representative examples of suchcounterions are the following: acetate, adipate, aminosalicylate,anhydromethylenecitrate, ascorbate, aspartate, benzoate,benzenesulfonate, bromide, citrate, camphorate, camphorsulfonate,chloride, estolate, ethanesulfonate, fumarate, glucoheptanoate,gluconate, glutamate, lactobionate, malate, maleate, mandelate,methanesulfonate, pantothenate, pectinate, phosphate/diphosphate,polygalacturonate, propionate, salicylate, stearate, succinate, sulfate,tartrate and tosylate. Other suitable anionic species will be apparentto the ordinarily skilled chemist.

[0135] Likewise, when more than one negative charge is necessary tomaintain charge neutrality, the counterion indicator may represent aspecie with more than one negative charge, such as malonate, tartrate orethylenediaminetetraacetate (EDTA), or two or more monovalent anions,such as chloride, etc. When a multivalent negatively charged counterionis present with a compound of formula I which bears a net singlepositive charge, an appropriate number of molar equivalents of theanionic species can be found in association therewith to maintain theoverall charge balance and neutrality.

[0136] Some of the compounds of formula I may be crystallized orrecrystallized from solvents such as organic solvents. In such casessolvates may be formed. This invention includes within its copestoichiometric solvates including hydrates as well as compoundscontaining variable amounts of solvents such as water that may beproduced by processes such as lyophilization. The compounds of formula Imay be prepared in crystalline form by for example dissolution of thecompound in water, preferably in the minimum quantity thereof, followedby admixing of this aqueous solution with a water miscible organicsolvent such as a lower aliphatic ketone such as a di-(C₁₋₆) alkylketone, or a (C₁₋₆) alcohol, such as acetone or ethanol.

[0137] A subset of compounds of formula I which is of interest relatesto those compounds where M¹ and M² are independently hydrogen ornegative charge, said negative charge(s) balanced by the appropriatenumber of counter balancing ions, and all other variables are asdescribed above.

[0138] Another subset of compounds of formula I which is of interestrelates to those compounds where R¹ and/or R² represents a C₁ to C₁₆straight, branched or unsaturated alkyl group substituted with 0 to 2R^(q), and substituted with 0 to 3 R_(x) groups and optionallyinterrupted by one of the following O, S, SO₂, —C(O)—, —C(O)—NR^(a)— and—CO₂-, provided that at least one of R¹ or R² contains an R^(q) and allother variables are described as above.

[0139] Another subset of compounds of formula I which is of interestrelates to those compounds where R¹ and/or R² represents C₄₋₁₂ straight,branched or unsaturated alkyl group optionally substituted with 1-2R_(x) and optionally substituted with 1-2 R^(q) groups, provided that atleast one of R¹ or R² contains an R^(q), wherein all other variables areas described above.

[0140] Another subset of compounds of formula I which is of interestrelates to those compounds where R¹ and/or R² represents a group of theformula:

[0141] wherein at least one R^(q) is present on R¹ or R² and all othervariables are defined as above.

[0142] Another subset of compounds of formula I which is of interestrelates to those compounds where R¹ and/or R² represents a group of theformula:

[0143] wherein at least one R^(q) group is present on R¹ or R² and allother variables are defined as above.

[0144] Another subset of compounds of formula I which is of interestrelates to those compounds where the relative and absolutestereochemistry is:

[0145] Still another subset of compounds of formula I which is ofinterest relates to those compounds where R¹ and/or R² represents agroup of the formula:

[0146] wherein A is (CH₂)₁₋₅ and

[0147] is phenyl, naphthyl, cyclohexyl or dibenzofuranyl, provided thatat least one of R¹ or R² contains an R^(q) and all other variables areas originally defined.

[0148] Still another subset of compounds of formula I that is ofinterest relates to those compounds where R¹ or R¹ represents a group ofthe formula:

[0149] wherein

[0150] A is (CH₂)₁₋₃, A′ is a single bond, —O— or (CH₂)₁₋₂

[0151] independently represent phenyl, thienyl, pyridyl, furanyl orcyclohexyl

[0152] Yet another subset of compounds of formula I, that is of interestrelates to those compounds where one of R¹ or R² is C₄₋₈ straight,branched or unsaturated alkyl optionally substituted with 1 to 2 R_(x)or a group of the formula:

[0153] where A is (CH₂)₁₋₂ and

[0154] is phenyl, cyclopentyl or cyclohexyl and the other of R¹ or R² is

[0155] i) a C₇₋₁₂ alkyl group substituted with R^(q),

[0156] ii) a group of the formula:

[0157]  where A is (CH₂)₁₋₂, A′ is a single bond,

[0158] is phenyl, thienyl or cyclohexyl and

[0159]  is phenyl, thienyl or pyridyl, or

[0160] iii) a group of the formula:

[0161]  where A is (CH₂ ₁₋₃,

[0162] is phenyl or thienyl and R^(q) is —(CH₂)₂₋₆-Q⁺Y⁻ and all othervariables are as originally defined.

[0163] Still another subset of compounds of formula I that is ofinterest relates to those compounds where:

[0164] R¹ is C₅₋₇ alkyl substituted with 0 to 2 R_(x) goups,

[0165] R² is C₇₋₁₀ alkyl substituted with 1R^(q) group and 0 to 2 R_(x)groups,

[0166]  and all other variables are as described above.

[0167] Still another subset of compounds of formula I that is ofinterest relates to those compounds where:

[0168] R¹ is:

[0169] R² is:

[0170]  and all other variables are as described above.

[0171] A preferred subset of R_(x) is R^(y).

[0172] It is preferred that a total of one or two R^(q) groups arepresent in R¹ and R² containing a total of 2 to 6 cationic nitrogenatoms. It is more preferred that a single R^(q) substituent is presentcontaining a tricationic or tetracationic Q⁺ group. A more preferredR^(q) is —E-Q⁺Y⁻ wherein E is (CH₂)₀₋₆ or —C(O)—N(R^(a))—(CH₂)₂₋₄, andQ⁺ is a tricationic or tetracationic group.

[0173] Preferred tricationic Q⁺ groups are:

[0174] wherein E¹ is (CH₂)₂₋₄ or —(CH₂)—C(O)—N(R^(a))—(CH₂)₂₋₄— andR^(a), Q¹ and Q² are as previously defined.

[0175] More preferred tricationic Q⁺ groups are:

[0176] wherein R^(a), R^(u), R^(v), and R^(w) are independently selectedas defined above.

[0177] Preferred tetracationic Q⁺ groups are:

[0178] wherein E¹ is (CH₂)₂₋₄ or —(CH₂)—C(O)—N(R^(a))—(CH₂)₂₋₄— andR^(a), Q¹, Q², and R^(w) are as defined above.

[0179] More preferred tetracationic Q⁺ groups are:

[0180] wherein R^(a), R^(u), R^(v), and R^(w) are as defined above.

[0181] Preferred. Y⁻ groups are chloride, bromide, acetate, citrate,succinate, phosphate, maleate, tartrate and sulfate.

[0182] The compounds of the invention, which are succinic acids orderivatives thereof, can be formulated in pharmaceutical compositions bycombining the compound with a pharmaceutically acceptable carrier.Examples of such carriers are set forth below. The compounds of formulaI have metallo-β-lactamase inhibitory properties, and are useful whencombined with a β-lactam antibiotic for the treatment of infections inanimals, especially mammals, including humans. The compounds may beused, for example, in the treatment of infections of, amongst others,the respiratory tract, urinary tract and soft tissues and blood.

[0183] The compounds may be employed in powder or crystalline form, inliquid solution, or in suspension. They may be administered by a varietyof means; those of principal interest include: topically, orally andparenterally by injection (intravenously or intramuscularly).

[0184] Compositions for injection, a preferred route of delivery, may beprepared in unit dosage form in ampules, or in multidose containers. Theinjectable compositions may take such forms as suspensions, solutions,or emulsions in oily or aqueous vehicles, and may contain variousformulating agents. Alternatively, the active ingredient may be inpowder (lyophillized or non-lyophillized) form for reconstitution at thetime of delivery with a suitable vehicle, such as sterile water. Ininjectable compositions, the carrier is typically comprised of sterilewater, saline or another injectable liquid, e.g., peanut oil forintramuscular injections. Also, various buffering agents, preservativesand the like can be included.

[0185] Topical applications may be formulated in carriers such ashydrophobic or hydrophilic bases to form ointments, creams, lotions, inaqueous, oleaginous or alcoholic liquids to form paints or in drydiluents to form powders.

[0186] Oral compositions may take such forms as tablets, capsules, oralsuspensions and oral solutions. The oral compositions may utilizecarriers such as conventional formulating agents, and may includesustained release properties as well as rapid delivery forms.

[0187] The compounds of the instant invention are metallo-β-lactamaseinhibitors, which are intended for use in pharmaceutical compositions.Accordingly, it is preferable that the metallo-β-lactamase inhibitorsare provided in substantially pure form, for example at least about 60%to about 75% pure, preferably about 85% to about 95% pure and mostpreferably about 98% or more pure (% are on a weight for weight basis).Impure preparations of the compounds may be used for preparing the morepure forms used in pharmaceutical compositions.

[0188] The dosage to be administered depends to a large extent upon thecondition and size of the subject being treated, the route and frequencyof administration, the sensitivity of the pathogen to the particularcompound selected, the virulence of the infection and other factors.Such matters, however, are left to the routine discretion of thephysician according to principles of treatment well known in theantibacterial arts. Another factor influencing the precise dosageregimen, apart from the nature of the infection and peculiar identity ofthe individual being treated, is the molecular weight of the compound.

[0189] The compositions for human delivery per unit dosage, whetherliquid or solid, may contain from about 0.01% to as high as about 99% ofactive material, the preferred range being from about 10-60%. Thecomposition will generally contain from about 15 mg to about 2.5 g ofthe active ingredient; however, in general, it is preferable to employdosage amounts in the range of from about 250 mg to 1000 mg. Inparenteral administration, the unit dosage will typically include thepure compound in sterile water solution or in the form of a solublepowder intended for solution, which can be adjusted to neutral pH andisotonic.

[0190] The invention described herein also includes a method of treatinga bacterial infection in a mammal in need of such treatment comprisingadministering to said mammal a compound of formula I in conjunction witha β-lactam antibiotic such as a carbapenem, penicillin or cephalosporinin an effective combination.

[0191] The preferred methods of administration of the Formula Icompounds include oral and parenteral, e.g., i.v., infusion, i.v. bolusand i.m. injection.

[0192] The compounds of formula I may suitably be administered to thepatient at a daily dosage of from 0.7 to 50 mg/kg of body weight. For anadult human (of approximately 70 kg body weight), from 50 to 3000 mg,preferably from 100 to 1000 mg, of a compound according to the inventionmay be administered daily, suitably in from 1 to 6, preferably from 2 to4, separate doses. Higher or lower dosages may, however, be used inaccordance with clinical practice.

[0193] The compounds may be used in combination with antibiotic agentsfor the treatment of infections caused by metallo-β-lactamase producingstrains, in addition to those infections which are subsumed within theantibacterial spectrum of the antibiotic agent. Metallo-β-lactamaseproducing strains include: Bacillus cereus, Bacteroides fragilis,Aeromonas hydrophila, Klebsiella pneumoniae, Pseudomonas aeruginosa,Serratia marcescens, Stenotrophomonas maltophilia, Shigella flexneri,Legionella gormanii, Chryseobacterium meningosepticum, Chryseobacteriumindologenes, Acinetobacter baumannii, Citrobacter freundii, andAeromonas veronii.

[0194] In accordance with the instant invention, it is generallyadvantageous to use a compound of formula I in admixture or conjuctionwith a carbapenem, penicillin, cephalosporin or other β-lactamantibiotic or prodrug. It also advantageous to use a compound of formulaI in combination with one or more β-lactam antibiotics, because of themetallo-β-lactamase inhibitory properties of the compounds. In thiscase, the compound of formula I and the β-lactam antibiotic can beadministered separately or in the form of a single compositioncontaining both active ingredients.

[0195] Carbapenems, penicillins, cephalosporins and other β-lactamantibiotics suitable for co-administration with the compounds of FormulaI, whether by separate administration or by inclusion in thecompositions according to the invention, include both those known toshow instability to or to be otherwise susceptible tometallo-β-lactamases and also known to have a degree of resistance tometallo-β-lactamase.

[0196] When the compounds of Formula I are combined with antibioticssuch as carbapenems dehydropeptidase (DHP) inhibitors may also becombined. Many carbapenems are susceptible to attack by a renal enzymeknown as DHP. This attack or degradation may reduce the efficacy of thecarbapenem antibacterial agent. Inhibitors of DHP and their use withcarbapenems are disclosed in, e.g., (European Patent 0007614, filed Jul.24, 1979 and application number 82107174.3, filed Aug. 9, 1982. Apreferred DHP inhibitor is7-(L-2-amino-2-carboxyethylthio)-2-(2,2-dimethylcyclopropanecarboxamide)-2-heptenoicacid or a useful salt thereof. Thus, compounds of the present inventionin combination with a carbapenem such as imipenem and a DHP inhibitorsuch as, cilastatin is contemplated within the scope of this invention.

[0197] A serine β-lactamase inhibitor such as clavulanic acid, sulbactamor tazobactam may also be co-administered with the compound of theinvention and β-lactam antibiotics, either by separate administration,or co-formulation with one, other or both of the compounds of theinvention and the β-lactam antibiotic.

[0198] Examples of carbapenems that may be co-administered with thecompounds of formula I include imipenem, meropenem, biapenem, (4R, 5S,6S)-3-[3S,5S)-5-(3-carboxyphenyl-carbamoyl)pyrrolidin-3-ylthio]-6-(1R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylicacid, (1S, 5R,6S)-2-(4-(2-(((carbamoylmethyl)-1,4-diazoniabicyclo[2.2.2]oct-1-yl)-ethyl(1,8-naphthosultam)methyl)-6-[1(R)-hydroxyethyl]-1-methylcarbapen-2-em-3-carboxylatechloride, BMS181139([4R-[4alpha,5beta,6beta(R*)]]-4-[2-[(aminoiminomethyl)amino]ethyl]-3-[(2-cyanoethyl)thio]-6-(1-hydroxyethyl)-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylicacid), BO2727 ([4R-3[3S*,5S*(R*)],4alpha,5beta,6beta(R*)]]-6-(1-hydroxyethyl)-3-[[5-[1-hydroxy-3-(methylamino)propyl]-3-pyrrolidinyl]thio]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylicacid monohydrochloride), E1010 ((1R, 5S,6S)-6-[1(R)-hydroxymethyl]-2-[2(S)-[1(R)-hydroxy-1-[pyrrolidin-3(R)-yl]methyl]pyrrolidin-4(S)-ylsulfanyl]-1-methyl-1-carba-2-penem-3-carboxylicacid hydrochloride), S4661((1R,5S,6S)-2-[(3S,5S)-5-(sulfamoylaminomethyl)pyrrolidin-3-yl]thio-6-[(1R)-1-hydroxyethyl]-1-methylcarbapen-2-em-3-carboxylicacid) and(1S,5R,6S)-1-methyl-2-{7-[4-(aminocarbonylmethyl)-1,4-diazoniabicyclo(2.2.2)octan-1yl]-methyl-fluoren-9-on-3-yl}-6-(1R-hydroxyethyl)-carbapen-2-em-3-carboxylatechloride.

[0199] Examples of penicillins suitable for co-administration with thecompounds according to the invention include benzylpenicillin,phenoxymethylpenicillin, carbenicillin, azidocillin, propicillin,ampicillin, amoxycillin, epicillin, ticarcillin, cyclacillin,pirbenicillin, azloccillin, mezlocillin, sulbenicillin, piperacillin,and other known penicillins. The penicillins may be used in the form ofpro-drugs thereof; for example as in vivo hydrolysable esters, forexample the acetoxymethyl, pivaloyloxymethyl, α-ethoxycarbonyloxy-ethyland phthalidyl esters of ampicillin, benzylpenicillin and amoxycillin;as aldehyde or ketone adducts of penicillins containing a6-α-aminoacetamido side chain (for example hetacillin, metampicillin andanalogous derivatives of amoxycillin); and as a-esters of carbenicillinand ticarcillin, for example the phenyl and indanyl α-esters.

[0200] Examples of cephalosporins that may be co-administered with thecompounds according to the invention include, cefatrizine,cephaloridine, cephalothin, cefazolin, cephalexin, cephacetrile,cephapirin, cephamandole nafate, cephradine, 4-hydroxycephalexin,cephaloglycin, cefoperazone, cefsulodin, ceftazidime, cefuroxime,cefmetazole, cefotaxime, ceftriaxone, and other known cephalosporins,all of which may be used in the form of pro-drugs thereof.

[0201] Examples of β-lactam antibiotics other than penicillins andcephalosporins that may be co-administered with the compounds accordingto the invention include aztreonam, latamoxef (Moxalactam-trade mark),and other known β-lactam antibiotics such as carbapenems like imipenem,meropenem or (4R, 5S,6S)-3-[(3S,5S)-5-(3-carboxyphenylcarbamoyl)pyrrolidin-3-ylthio]-6-(1R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylicacid, all of which may be used in the form of pro-drugs thereof

[0202] Preferred carbapenems are imipenem, meropenem and (4R, 5S,6S)-3-[(3S,5S)-5-(3-carboxyphenylcarbamoyl)pyrrolidin-3-ylthio]-6-(1R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylicacid.

[0203] Particularly suitable penicillins for co-administration with thecompounds according to the invention include ampicillin, amoxycillin,carbenicillin, piperacillin, azlocillin, mezlocillin, and ticarcillin.Such penicillins may be used in the form of their pharmaceuticallyacceptable salts, for example their sodium salts. Alternatively,ampicillin or amoxycillin may be used in the form of fine particles ofthe zwitterionic form (generally as ampicillin trihydrate or amoxycillintrihydrate) for use in an injectable or infusable suspension, forexample, in the manner described herein in relation to the compounds offormula I. Amoxycillin, for example in the form of its sodium salt orthe trihydrate, is particularly preferred for use in compositionsaccording to the invention.

[0204] Particularly suitable cephalosporins for co-administration withthe compounds according to the invention include cefotaxime, ceftriaxoneand ceftazidime, which may be used in the form of their pharmaceuticallyacceptable salts, for example their sodium salts.

[0205] When the compositions according to this invention are presentedin unit dosage form, each unit dose may suitably comprise from about 25to about 1000 mg, preferably about from 50 to about 500 mg, of acompound according to the invention. Each unit dose may, for example, be62.5, 100, 125, 150, 200 or 250 mg of a compound according to theinvention.

[0206] When the compounds of formula I are co-administered with apenicillin, cephalosporin, carbapenem or other β-lactam antibiotic, theratio of the amount of the compounds of formula I to the amount of theother β-lactam antibiotic may vary within a wide range. The said ratiomay, for example, be from 100:1 to 1:100; more particularly, it may forexample, be from 2:1 to 1:30. The amount of carbapenem, penicillin,cephalosporin or other β-lactam antibiotic according to the inventionwill normally be approximately similar to the amount in which it isconventionally used.

[0207] The claimed invention also includes the use of a compound offormula I, a pharmaceutically acceptable salt, ester, prodrug, anhydrideor solvate thereof, in the manufacture of a medicament for the treatmentof bacterial infections.

[0208] The claimed invention also includes the use of a compound offormula I as a metallo-β-lactamase inhibitor.

[0209] The claimed invention further includes a method of treatingbacterial infections in humans or animals which comprises administering,in combination with a β-lactam antibiotic, a therapeutically effectiveamount of a metallo-β-lactamase inhibitor of formula I.

[0210] The claimed invention further includes a method of treatingbacterial infections in humans or animals which comprises administering,in combination with a carbapenem antibiotic, a therapeutically effectiveamount of a metallo-β-lactamase inhibitor of formula I.

[0211] The claimed invention also includes a composition comprising ametallo-β-lactamase inhibitor of formula I together with a β-lactamantibiotic and a pharmaceutically acceptable carrier.

[0212] The claimed invention also includes a composition comprising ametallo-β-lactamase inhibitor of formula I together with a carbapenemantibiotic and a pharmaceutically acceptable carrier.

[0213] The compositions discussed above may optionally include a serineβ-lactamase inhibitor as described above as well as a DHP inhibitor.

[0214] Using standard susceptibility tests the compounds of the instantinvention were found to be active against metallo-β-lactamase enzymesproduced by a range of organisms.

[0215] The compounds of the present invention are synthesized using thegeneral conditions shown in the accompanying flow charts (A through F).

[0216] The 2,3-disubstituted succinic acid compounds of the presentinvention can be prepared as described in Flow Sheets A-F. The cationicR^(q) substituents of the compounds of the present invention aregenerally carried through the syntheses in protected or precursory formand are then deprotected or elaborated near or at the end of thesynthesis. Introduction of the R^(q) substituent from a precursor groupis described in detail in Flow Sheet F.

[0217] The synthesis of Flow Sheet A is based on a known literatureprocedure (M. J. Crimmin et. al., SynLett 1993, 137). Referring to FlowSheet A, the R¹-substituted acetic acid starting materials A1 arereadily available from commercial sources or are readily prepared by avariety of methods known in the art. Briefly, the starting material A1is alkylated with an ester derivative of bromoacetic acid, employing achiral auxiliary group to achieve stereoselectivity in the reaction.After removal of the chiral auxiliary to give A4, the R²* group isintroduced stereoselectively by an alkylation reaction to give A5. TheR²* group may be R² as defined above, or may contain R^(x) and R^(q)substituents in precursory or protected form which require elaboration.Such elaboration may be carried-out at this point. Removal of thecarboxyl protecting group of A5 then provides the final compound A6.

[0218] The first step of Flow Sheet A is introduction of the chiralauxiliary. A suggested method is as follows. A mixed anhydride is formedbetween the starting carboxylic acid A1 and pivalic acid by treating A1with a tertiary amine base such as triethylamine and pivaloyl chloridein a suitable ethereal solvent such as tetrahydrofuran at reducedtemperature such as between −78° C. and 0° C. After a suitable reactiontime, such as from 30 min to 3 hours, the resulting activatedintermediate is then reacted with a freshly prepared solution oflithio-(4R)-benzyl-2-oxazolidinone in tetrahydrofuran at reducedtemperature such as between −78° C. and 0° C. After conventionalisolation and purification, intermediate A2 is obtained. Intermediate A2is deprotonated with a strong base such as sodium hexamethyldisilazidein a solvent such as tetrahydrofuran at reduced temperature such asbetween −78° C. and −70° C. The resulting enolate is alkylated byaddition of BrCH₂CO₂P¹, where P¹ is a removable carboxyl protectinggroup. After an appropriate reaction period, such as from 1 to 3 hours,compound A3 is obtained by conventional isolation and purificationtechniques. Suitable removable ester derivatives of bromoacetic acid forthis alkylation reaction are t-butyl bromoacetate, allyl bromoacetate orbenzyl bromoacetate.

[0219] The oxazolidinone chiral auxiliary group of A3 is removed by ahydrolysis reaction. Aqueous lithium hydroxide and aqueous hydrogenperoxide are employed for this reaction along with an organic co-solventsuch as tetrahydrofuran. The reaction is carried-out at a temperature offrom 0° C. to 30° C. for a reaction time of from 30 min to 4 hours.After acidification, conventional isolation and purification providesintermediate A4.

[0220] An alternative method of removing the chiral auxilliary consistsof reacting A3 with lithium benzyloxide (LiOCH₂Ph) followed by cleavageof the resulting benzyl ester to give A4. The reaction of A3 withlithium benzyloxide is carried-out in tetrahydrofuran as solvent at atemperature of from −78° C. to 30° C. for a reaction time of from 30 minto 4 hours. Cleavage of the resulting benzyl ester is accomplished inconventional fashion, eg by hydrogenolysis employing a suitable catalystsuch as palladium on carbon in an appropriate solvent such as ethanol at1-2 atmospheres pressure of hydrogen. After conventional isolation andpurification, compound A4 is obtained.

[0221] Alkylation of A4 to give A5 is accomplished by deprotonating A4with >2 equivalents of a strong hindered base to give a dianion which isthen reacted with an alkylating agent R²*L to give A5, where R²* is asdefined above and L is a displaceable leaving group such as iodide,bromide or trifluoromethanesulfonate. The reaction proceeds with highstereoselectivity to give predominately the stereoisomer shown in FlowSheet A. The deprotonation reaction is carried-out in a suitable solventsuch as tetrahydrofuran at a temperature of from −78° C. to −70° C. fora reaction time of from 30 min to 3 hours. Preferred bases for thisreaction are lithium bis(trimethylsilyl)amide and lithiumdiisopropylamide. After addition of the alkylating agent, the reactionis allowed to proceed at a temperature of from −78° C. to 25° C. for areaction time of from 1 to 12 hours. Progress of the reaction can bemonitored by conventional analytical methods, eg HPLC and TLC. Preferredalkylating agents for this reaction are alkyl iodides and alkylbromides. Other suitable alkylating agents are well known in the art andinclude alkyl trifluoromethanesulfonates, alkyl methanesulfonates andalkyl tosylates. After conventional isolation and purification,intermediate A5 is obtained. The minor stereoisomer produced in thisreaction can often be separated from A5 at this stage by conventionalchromatographic techniques. However, it is often preferable to carry-outthis separation at the stage of A6, after removal of the carboxylprotecting group as described below.

[0222] Deprotection of any R^(x) or R^(q) groups which are present inprotected form may be accomplished at this point. For example, ifcompound A5 contains a protected hydroxyl or amino group, saidprotecting group may conveniently be removed at the stage of A5.Alternatively, depending on the nature of the protecting group it may beremoved concurrent with or subsequent to the removal of the. carboxylprotecting group as described immediately below. Introduction of thecationic R^(q) group may also be accomplished at this point from aprecursor substituent. This procedure is described in detail in FlowSheet F further below.

[0223] Removal of the carboxyl protecting group of A5 by standardmethods gives the final compound A6. When P¹ is t-butyl, this isaccomplished by treating A5 with a strong acid such as trifluoroaceticacid in a suitable solvent such as dichloromethane. The reaction iscarried-out at a temperature of from 0° C. to 30° C. for a reaction timeof from 1 to 8 hours. The final compound A6 is then isolated byconventional techniques. Other methods of removing tert-butyl estergroups are known in the art and may also be employed (see e.g. Greene,T. W., et al. Protective Groups in Organic Synthesis, John Wiley & Sons.Inc., 1991).

[0224] It will be apparent to one skilled in the art that employing achiral auxiliary of the opposite absolute configuration [eg.lithio-(4S)-benzyl-2-oxazolidinone] in the first step of Flow Sheet Awill make possible the synthesis of compound A3 with the alternativestereochemistry at the newly created stereocenter. This will in turnmake possible the synthesis of the final compounds A6 of Flow Sheet A,with the opposite absolute configuration. Other chiral auxiliary groupsare also known in the art and may also be employed.

[0225] Flow Sheet B illustrates a variation of Flow Sheet A which ispreferred in certain cases, for example when Ar¹ is a heteroaryl groupsuch as pyridyl. In this synthesis the second substituent on thesuccinic acid is introduced by an aldol reaction instead of analkylation reaction. The synthesis begins with compound A4, which isprepared as described in Flow Sheet A. Compound A4 is deprotonatedwith >2 equivalents of a strong hindered base to give a dianion which isthen reacted with an aldehyde Ar¹CHO to give B1, where Ar¹ is anoptionally substituted aryl or heteroaryl group, terms which are definedabove. The deprotonation reaction is carried-out in a suitable solventsuch as tetrahydrofuran at a temperature of from −78° C. to −70° C. fora reaction time of from 30 min to 3 hours. Preferred bases for thisreaction are lithium bis(trimethylsilyl)amide and lithiumdiisopropylamide. After addition of the aldehyde, the reaction isallowed to proceed at a temperature of from −78° C. to 25° C. for areaction time of from 1 to 12 hours. After conventional isolation andpurification, intermediate B1 is obtained.

[0226] Compound B1 is next cyclized to the lactone B2. Suitableconditions for this cyclization reaction would be exposure of B1 toacetic anhydride and triethylamine in an inert solvent such asdichloromethane. Reductive opening of lactone B2, such as byhydrogenolysis over palladium on carbon in a-suitable solvent such asmethanol, provides compound B3. Deprotection of any R^(x) or R^(q)groups which are present in protected form may be accomplished at thispoint. In addition, introduction of a cationic R^(q) group from aprecursor substituent may be carried-out at, the stage of B3. Thisprocedure is described in detail in Flow Sheet F further below. Removalof the carboxyl protecting group of B3 by conventional methods thengives the final compound B4.

[0227] Flow Sheet C illustrates an extension of the synthesis of FlowSheet A which makes possible the introduction of a variety of preferredbiaryl-type R² substituents. Briefly, starting with compound A4 fromFlow Sheet A, alkylation with K—Ar²—(CH₂)_(n)—L by the method describedin Flow Sheet A gives intermediate C1; where L is a displaceable leavinggroup such as iodide, bromide or trifluoromethanesulfonate, n is 1,2,3or 4, Ar2 is an optionally substituted aryl or heteroaryl group asdefined above, and K is iodide, bromide, chloride or a protectedhydroxyl group which can be converted to a trifluoromethanesulfonategroup by known methods. Protection of the free carboxyl group of C1 witha removable protecting group p² gives C2. When K is a protected hydroxylgroup it is deprotected and converted to a trifluoromethanesulfonategroup at this point. A palladium catalyzed organometallic cross-couplingreaction between C2 and an organometallic reagent R³-Met gives compoundC3; where Met is a boronic acid or trialkyltin moiety and R³ is anoptionally substituted alkenyl, alkynyl, aryl or heteroaryl group asdefined above. Deprotection or elaboration of any R^(x) or R^(q) groupswhich are present in protected or precursory form is accomplished atthis point. Removal of the two carboxyl protecting groups of C3 thenprovides the final compound C4.

[0228] The p² carboxyl protecting group is introduced in conventionalfashion. A preferred p² group is p-methoxybenzyl which can be introducedemploying p-methoxybenzyl alcohol, a carbodiimide reagent such as1,3-diisopropylcarbodiimide and N,N-dimethylaminopyridine catalyst in asuitable inert solvent such as dichloromethane. Other suitable esterprotecting groups known in the art could also be employed (see e.g.Greene, T. W., et al. Protective Groups in Organic Synthesis, John Wiley& Sons. Inc., 1991).

[0229] The palladium catalyzed cross-coupling reaction between C2 andR³-Met is carried-out by procedures known in the scientific and patentliterature. When Met is a boronic acid moiety [—B(OH)₂] the reaction iscommonly known as a Suzuki reaction (see Suzuki, Chem. Rev. 1995, 95,2457). Compound C2 is combined with the boronic acid R³—B(OH)₂ in acoupling solvent such as 1,2-dimethoxyethane, N,N-dimethylformamide ortoluene, optionally with water as a co-solvent, with a base such assodium carbonate and a palladium catalyst such astetrakis(triphenylphosphine)-palladium(0). The reaction is carried-outat a temperature of from 20° C. to 125° C. for a reaction time of from 1to 48 hours. The coupled product C3 is then isolated by conventionaltechniques. When Met is a trialkyltin moiety, the reaction is commonlyknown as a Stille reaction and the cross-coupling is carried-out byprocedures well known in the literature (T. N. Mitchell, Synthesis 1992,803).

[0230] Deprotection of any R^(x) or R^(q) groups which are present inprotected form may be accomplished at this point. In addition,introduction of a cationic R^(q) group from a precursor substituent maybe carried-out at the stage of C3. This procedure is described in detailin Flow Sheet F further below.

[0231] Removal of the carboxyl protecting groups of C3 by standardmethods provides the final compound C4. It is often convenient for theprotecting groups p¹ and p² to be selected such that they can both beremoved under the same reaction conditions. For example, when p¹ istert-butyl and p² is p-methoxybenzyl, both esters of C3 can be removedin a single step by treating C3 with a strong acid such astrifluoroacetic acid in a suitable solvent such as dichloromethane. Itis sometimes advantageous to include a trapping agent such astriethylsilane or anisole in the reaction mixture. The reaction iscarried-out at a temperature of from 0° C. to 30° C. for a reaction timeof from 1 to 8 hours. The final compound C4 is then isolated byconventional techniques. Other methods of removing tert-butyl andp-methoxybenzyl ester groups are known in the art and may also beemployed (see e.g. Greene, T. W., et al. Protective Groups in OrganicSynthesis, John Wiley & Sons. Inc., 1991). Flow Sheet D illustrates analternative synthesis of compounds of the present invention. TheR¹-substituted acetic acid starting materials D1 (M=H) and theesterified derivatives thereof (M=esterifying group) are readilyavailable from commercial sources or are readily prepared by a varietyof methods known in the art. The synthesis of Flow Sheet D is based onknown literature procedures (see for example J. L. Belletire and D. F.Fry, J. Org. Chem. 1987, 52, 2549). Briefly, starting material D1 isdeprotonated with a strong base and the resulting dianion (M=H) or anion(M=esterifying group) is oxidatively coupled with a suitable oxidizingreagent. In the case of M=H, acidic work-up and conventional isolationand purification gives the final compound D2. In the case ofM=esterifying group, an additional deprotection step is also needed. Apreferred strong base for the deprotonation reaction is lithiumdiisopropylamide. Suitable oxidizing agents for the synthesis of FlowSheet D include iodine, copper(II) salts such as CuBr2, and titaniumtetrachloride.

[0232] In the synthesis of Flow Sheet D, protection or elaboration ofany R^(x) or R^(q) groups which are present in protected or precursoryform is best accomplished where M=esterifying group, prior to removal ofsaid estenrfying group. In addition, introduction of a cationic R^(q)group from a precursor substituent may be carried-out as described indetail in Flow Sheet F further below.

[0233] Since the synthesis of Flow Sheet D is based on adimerization-type reaction, it is best suited for the synthesis ofsymmetrically 2,3-disubstituted succinic acids (R¹=R²). For this reason,it is generally less preferred than the syntheses of Flow Sheets A, Band C. The synthesis of Flow Sheet D also generally produces a racemicmixture of stereoisomers., However, it is possible to employ a chiralauxiliary in the synthesis of Flow Sheet D in order to achieve highstereoselectivity and optical purity (see for example N. Kise et. al. J.Org. Chem. 1995, 60, 1100). Such use of a chiral auxiliary isillustrated in Flow Sheet E.

[0234] Flow Sheet F illustrates a suggested method for the introductionof the cationic substituents of the compounds of the present inventionfrom a precursor substituent. The starting material F1 of Flow Sheet Fis substituted with a precursor substituent which can be elaborated intothe desired cationic substitutent, R^(q). A preferred precursorsubstituent is a protected hydroxymethyl group, P³OCH₂—, where p³ is aremovable hydroxyl protecting group. In Flow Sheet F, R⁴ is defined suchthat the moiety [—R⁴—CH₂—Q⁺Y⁻] represents an R² group as defined above.Examples of representative R⁴ groups are shown in Table 1.

[0235] The starting material F1 of Flow Sheet F is synthesized by onethe methods described in Flow Sheets A, B, C, D, and E. When F1 issynthesized according to Flow Sheet A, it is derived from intermediateA5, through protection of the free carboxyl group with an appropriatecarboxyl protecting group p². In this case, the precursor substituent ispresent in the R¹ or R²* substituent of A5. When F1 is synthesizedaccording to Flow Sheet B, it is derived from intermediate B3, throughprotection of the free carboxyl group with an appropriate carboxylprotecting group p². In this case, the precursor substituent is presentin the R¹ or Ar¹ substituent of B3. When F1 is synthesized according toFlow Sheet C, it is derived from or corresponds to, intermediate C3. Inthis case, the precursor substituent is present in the R¹, Ar² or R³substituent of C3. Starting material F1 may also be prepared byappropriate modification of the syntheses of Flow Sheets D and E aswould be apparent to those skilled in the art.

[0236] Referring to Flow Sheet F, the first step is removal of thehydroxyl protecting group p³. This is accomplished by conventionalmethods. Hydroxyl protecting group p³ is generally selected such that itmay be selectively removed in the presence of the carboxyl protectinggroups P¹ and p² A preferred p³ is t-butyldimethylsilyl. Removal of thepreferred t-butyldimethylsilyl p³ is accomplished by treating F1 withtetra-n-butylammonium fluoride and acetic acid in tetrahydrofuran assolvent. Other hydroxyl protecting groups are well known in the art andmay also be employed (see e.g. Greene, T. W., et al. Protective Groupsin Organic Synthesis, John Wiley & Sons. Inc., 1991).

[0237] Introduction of the cationic substituent is accomplished by anactivation-dispacement process. Briefly, the hydroxyl group of F2 isconverted into a suitable leaving group, G, which is thereafterdisplaced with a nucleophilic nitrogen. compound Q*, to yield F4. Withcertain Q* groups, additional steps may also be needed such as removalof amino protecting groups or conversion of an amine precursor suchazide into an amino group. The protecting groups are removed from F4 inconventional fashion and then in an optional step a pharmaceuticallyacceptable counterion Y⁻ may be introduced to provide compound F5.

[0238] The following are examples of suitable leaving groups G: alkyland substituted alkylsulfonates, aryl and substituted arylsulfonates andhalides. The common sulfonate leaving groups are: methanesulfonyloxy,trifluoromethanesulfonyloxy, fluorosulfonyloxy, p-toluenesulfonyloxy,and 2,4,6-triisopropylbenzenesulfonyloxy. The preferred halogen leavinggroups are bromide and iodide.

[0239] Compound Q* represents a precursor group to the cationic group Q⁺as defined above. As such, it may require further modification after itsreaction with F3. The nucleophilic nitrogen moiety of Q* is generallythe nitrogen of a primary, secondary or tertiary amino group or a ringnitrogen of a heteroaryl group such as a 1-substituted-imidazole. Inaddition to its nucleophilic nitrogen atom, Q* may include 1,2 or 3 ofthe following moieties: positively charged nitrogen atoms, protectedamino groups, amine precursor groups such as azido. Suitable protectinggroups for amino groups present in Q* would be t-butyloxycarbonyl-,allyloxycarbonyl- and p-nitrobenzyloxycarbonyl-. The Q* groups may beprepared by standard methods known in the scientific and patentliterature. Suitable Q* groups are listed in Table 2.

[0240] Referring to Flow Sheet F, the hydroxyl group of F2 may beconverted into a suitable alkyl- or arylsulfonate leaving group bytreating with an appropriate agent such as an alkyl- or arylsulfonylchloride or an alkyl- or arylsulfonic anhydride in the presence of ahindered organic base such as triethylamine or 2,6-lutidine. A suitablesolvent such as dichloromethane is employed and the reaction is carriedout at reduced temperature, such as from about −70° C. to 0° C.

[0241] The preferred halogen leaving groups may be introduced bydisplacing an alkyl- or arylsulfonate leaving group with an appropriatemetal halide. Thus, compound F3, where G is an alkyl- or arylsulfonategroup, is reacted with a suitable metal halide such as sodium iodide orpotassium bromide in a suitable solvent such as acetone, acetonitrile,tetrahydrofuran, 1-methyl-2-pyrrolidinone and the like, at from about 0°C. to 50° C. Alternatively, the hydroxyl group of F2 may be directlyconverted into an iodide group by reaction with an appropriate reagent,eg. by treatment of F2 with methyl triphenoxyphosphonium iodide in asuitable solvent, such as N,N-dimethylformamide, at reduced or ambienttemperatures. Introduction of the cationic substituent is accomplishedby reacting F3 with a nucleophilic nitrogen compound Q* in a suitablesolvent, such as acetonitrile, tetrahydrofuran, 1-methyl-2-pyrrolidinoneand the like, at about 0° C. to 50° C. to provide F4. When the leavinggroup, G, is iodide or bromide, this displacement reaction may also befacilitated by the addition of silver trifluoromethanesulfonate to thereaction mixture.

[0242] When the hydroxyl group of F2 is located at a benzylic position,and the reactive trifluoromethanesulfonate group is employed as theleaving group G in F3, the activation and displacement steps must becarried-out in situ, since in this case F3 cannot be isolated byconventional techniques due to its instability. Thus, treatment of F2with a slight excess of trifluoromethanesulfonic anhydride in thepresence of a hindered, non-nucleophilic base such as 2,6-lutidine,2,4,6-collidine, or 2,6-di-tert-butyl-4-methyl-pyridine in a suitablesolvent, such as dichloromethane or acetonitrile, at from about −78° C.to −20° C. provides for the generation of the trifluoromethanesulfonateactivating group. Introduction of the cationic group is thenaccomplished by reacting the above trifluoromethanesulfonateintermediate in situ with Q* at reduced temperature. It is also possiblein certain instances to use the nucleophilic nitrogen compound Q* as thebase for the formation of the trifluoromethanesulfonate activatinggroup. In this case, treatment of F2 with trifluoromethanesulfonicanhydride in the presence of at least two equivalents of Q* at reducedtemperature such as from −78° C. to 0° C. provides intermediate F4.Examples of Q* which are suitable for use in this manner are1-methylimidazole and 1,4-diazabicyclo(2.2.2)octane.

[0243] Removal of the carboxyl protecting groups of F4 by standardmethods provides the final compound F5. If Q* includes one or moreprotected amino groups, these are removed either before, after orsimultaneous with the carboxyl protecting groups depending on the exactnature of the protecting groups. If Q* includes one or more amineprecursor groups, these may be converted to the desired amine or amineseither before or after removal of the carboxyl protecting groupsdepending on the nature of the protecting groups. In the case of anazido amine precursor group, this may be accomplished by hydrogenationover a suitable catalyst such as rhodium on carbon. After the protectinggroups are removed from F4, and the cationic group Q⁺ has been fullyelaborated, the final compound F5 is isolated by conventionaltechniques. As an optional final step, a pharmaceutically acceptablecounterion Y⁻, which may differ from G⁻, may be introduced by standardtechniques, e.g. by employing an anion exchange resin. Suitablenegatively charged counterions are listed above under the description ofpharmaceutically acceptable salts.

[0244] Compound F5 is electronically balanced. If more than one positivecharge is present in the cationic Q⁺ group of F5, it is understood thatan appropriate amount of negative counterion is Y⁻ present to result inoverall electronic balance in the final compound F5. Likewise, it isunderstood that when the counterion Y⁻ is an anionic species possessingmore than one negative charge, then an appropriate amount of Y⁻ ispresent to result in overall electronic balance in the final compound ofFormula I. For example, when Y⁻ is a dianionic species, then one-half ofa molar equivalent of Y⁻ is present relative to the succinate moiety.

[0245] Representative examples of R⁴ and Q* are found below in Tables 1and 2 respectively: TABLE 1 Representative P³OCH₂—R⁴ Groups

[0246] TABLE 2 Representative Q* Groups

[0247] The invention is further described in connection with thefollowing non-limiting examples.

PREPARATION 1

[0248]

[0249] A solution of (4R)-benzyl-2-oxazolidinone (2.44 g, 13.77 mmol) in100 mL of THF was cooled to −70° C. and metalated by the dropwiseaddition of a 2.5M solution of n-butyllithium in hexanes (5.52 mL, 13.77mmol). After 20 min, neat hydrocinnamoyl chloride (2.05 ml, 13.79 mmol)was added. After 15 min, the reaction mixture was warmed by placing inan ice bath and kept at 0° C. C for 1 hr. The reaction was hydrolyzed bythe addition of sat. aqueous NH₄Cl and most of the THF was removed byrotary evaporation. The residue was partitioned between ethyl acetateand sat. aqueous NH₄Cl and the organic phase was washed with sat.aqueous NaHCO₃, water and brine. The organic layer was dried over Na₂SO₄and evaporated in vacuo to give a solid. Flash chromatography through500 g of silica gel (50:40:10 hexane/CH₂Cl₂/EtOAc) yielded 3.89 g of thetitle compound as a white solid.

[0250]¹H-NMR (500 Mz, CDCl₃): δ 2.79 (dd, J=13.5, 9.4 Hz, 1H), 3.02-3.13(m, 2H), 3.24-3.41 (m, 3H), 4.16-4.21 (m, 2H), 4.65-4.74 (m, 1H),7.16-7.40 (m, 10H). MS (CI): m/z=385.2 (MH⁺).

PREPARATION 2

[0251]

[0252] A stirred solution of compound 1 (3.283 g, 10.612 mmol) in 35 mLof THF was cooled to −78° C. and a 1.0M solution of NaN(TMS)₂ in THF(11.67 mL, 11.67 mmol) was added dropwise during 15 min. After 30 min, asolution of t-butyl bromoacetate (2.04 mL, 13.82 mmol) in 2 mL of THFwas added dropwise during 5 min. The solution was stirred at −78° C. for1 h and then the ice bath was removed and stirring was continued for 1h. The reaction was hydrolyzed by the addition of sat. aqueous NH4Cl andmost of the THF was removed by rotary evaporation. The residue waspartitioned between ethyl acetate and sat. aqueous NH4Cl and the organicphase was washed with water and brine. The organic layer was dried overNa2SO₄ and evaporated in vacuo to give a solid. Flash chromatographythrough 410 g of silica gel (35:60:5 hexane/CH₂Cl₂/EtOAc) yielded 2.86 gof the title compound as a white foam.

[0253]¹H-NMR (500 Mz, CDCl₃): δ 1.43 (s, 9H), 2.41 (dd, J=17.0, 4.1 Hz,1H), 2.64-2.80 (m, 2H), 2.88 (dd, J=17.0, 11.0 Hz, 1H), 3.04 (dd,J=13.0, 6.3 Hz, 1H), 3.34 (dd, J=13.5, 3.2 Hz, 1H), 3.95 (t, J=8.4 Hz,1H), 4.08-4.12 (m, 1H), 4.5-4.6 (m, 2H), 7.21-7.40 (m, 10H). MS (ESI):m/z=441.3 (M+NH₄ ⁺).

PREPARATION 3

[0254]

[0255] A stirred solution of compound 2 (2.860 g, 6.753 mmol) in 70 mLof 4:1 THF/H₂O was cooled to 0° C. and 30% aq. hydrogen peroxide (2.8mL, 27.01 mmol) was added dropwise during 5 min. After 5 min, a 1.0Msolution of LiOH.H₂O in H₂O (13.51 ml, 13.51 mmol) was added dropwiseduring 10 min. The reaction was kept at 0° C. for 1.75 hr. and then a1.5M solution of Na₂SO₃ in H₂O (18.0 ml, 27.01 mmol) was added. The icebath was removed and the reaction mixture was allowed to warm towardsroom temperature over 30 min. A solution of 1.0N NaHCO₃ in H₂O was addeduntil the reaction mixture had a pH=9 by pH paper (˜5 ml). Most of theTHF was removed by rotary evaporation and the residue was partitionedbetween CH₂Cl₂ and H₂O. The aqueous layer was washed 3×CH₂Cl₂ and thenacidified with 2N HCl until pH=3 by pH paper. The aqueous layer wasextracted 4×CH₂Cl₂ and the combined organic extracts were dried overNa2SO4 and evaporated in vacuo to give an oil. Flash chromatographythrough 100 g of silica gel (94:6 CH₂Cl₂/MeOH+0.5% HOAc) yielded 1.74 gof the title compound as a white solid.

[0256]¹H-NMR (500 Mz, CDCl₃): δ 1.45 (s, 9H), 2.38 (dd, J=16.6, 4.4 Hz,1H), 2.58 (dd, J=16.6, 8.5 Hz, 1H), 2.80 (dd, J=15.3, 10.3 Hz, 1H),3.10-3.20 (m, 2H), 7.20-7.40 (m, 5H), 11.99 (bs, 1H). MS (ESI):m/z=430.2 (M+NH₄ ⁺).

PREPARATION 4

[0257]

[0258] To a stirred solution of 3-(4-biphenyl)-propionic acid (1.805 g,7.977 mmol) in 40 mL of THF was added Et₃N (1.28 mL, 9.17 mmol) and thesolution was cooled to −70° C. Neat pivaloyl chloride (1.0 ml, 8.1 mmol)was added and a thick white suspension resulted. After 15 min, thereaction mixture was warmed by placing in an ice bath and kept at 0° C.for 40 min. The mixture was then re-cooled to −70° C. In a separateflask, a solution of (4R)-benzyl-2-oxazolidinone (1.44 g, 8.13 mmol) in35 mL of THF was cooled to −70° C. and metalated by the dropwiseaddition of a 2.5M solution of n-butyllithium in hexanes (3.25 mL, 8.13mmol). The resulting anion solution was added to the re-cooledsuspension via a cannula, rinsing with an additional 3 mL of THF. After15 min, the reaction mixture was warmed by placing in an ice bath andkept at 0° C. for 30 min. The reaction was hydrolyzed by the addition ofsat. aqueous NH₄Cl and most of the THF was removed by rotaryevaporation. The residue was partitioned between ethyl acetate and sat.aqueous NH₄Cl and the organic phase was washed with sat. aqueous NaHCO₃,water and brine. The organic layer was dried over Na₂SO₄ and evaporatedin vacuo to give a solid. Flash chromatography through 240 g of silicagel (CH₂Cl₂) yielded 2.45 g of the title compound as a white solid.

[0259]¹H-NMR (500 Mz, CDCl₃): δ 2.79 (dd, J=13.3, 9.4 Hz, 1H), 3.05-3.15(m, 2H), 3.25-3.41 (m, 3H), 4.15-4.25 (m, 2H), 4.65-4.75 (m, 1H),7.15-7.65 (m, 14H). MS (EI): m/z=385.2 (M⁺).

PREPARATION 5

[0260]

[0261] A stirred solution of compound 4 (1.000 g, 2.594 mmol) in 40 mLof THF was cooled to −78° C. and a 1.0M solution of NaN(TMS)₂ in THF(2.85 mL, 2.85 mmol) was added dropwise during 5 min. After 30 min, asolution of t-butyl bromoacetate (0.500 mL, 3.37 mmol) in 4 mL of THFwas added dropwise during 5 min. The solution was stirred at −78° C. for1 h and then the reaction was hydrolyzed by the addition of sat. aqueousNH₄Cl. The reaction mixture was partitioned between ethyl acetate andsat. aqueous NH₄Cl and the organic phase was washed with water andbrine. The organic layer was dried over Na₂SO₄ and evaporated in vacuoto give a solid. Flash chromatography through 150 g of silica gel(65:30:5 hexane/CH₂Cl₂/EtOAc) yielded 1.12 g of the title compound as awhite solid.

[0262]¹H-NMR (500 Mz, CDCl₃): δ 1.43 (s, 9H), 2.45 (dd, J=16.9, 4.1 Hz,1H), 2.65-2.80 (m, 2H), 2.89 (dd, J=16.9, 10.8 Hz, 1H), 3.07 (dd,J=13.0, 6.1 Hz, 1H), 3.34 (dd, J =13.5, 3.0 Hz, 1H), 3.94 (t, J=8.4 Hz,1H), 4.08-4.11 (m, 1H), 4.5-4.6 (m, 2H), 7.25-7.60 (m, 14H). MS (ESI):m/z=517.5 (M+NH₄ ⁺).

PREPARATION 6

[0263]

[0264] A stirred solution of compound 5 (0.907 g, 1.815 mmol) in 10 mLof THF was cooled to −70° C. and a freshly prepared 0.27 M solution ofLiOBn in THF (10 mL, 2.7 mmol) was added dropwise during 10 min. Thereaction was allowed to warm gradually to −10° C. during 2 h and wasthen placed in an ice bath and kept at 0° C. for 50 min. The reactionmixture was partitioned between ethyl acetate and sat. aqueous NH₄Cl andthe organic phase was washed with water and brine. The organic layer wasdried over Na₂SO₄ and evaporated in vacuo to give an oil. Flashchromatography through 125 g of silica gel (75:20:5 hexane/CH₂Cl₂/EtOAc)yielded 0.696 g of the title compound as a white solid.

[0265]¹H-NMR (500 Mz, CDCl₃): δ 1.42 (s, 9H), 2.43 (dd, J=16.6, 5.1 Hz,1H), 2.67 (dd, J=16.6, 9.1 Hz, 1H), 2.85 (dd, J=13.6, 7.9 Hz, 1H), 3.07(dd, J=13.5, 6.9 Hz, 1H), 3.15-3.25 (m, 1H), 5.09 (d, J=12.4 Hz, 1H),5.15 (d, J=12.4 Hz, 1H), 7.20-7.65(m, 14H). MS (EI): m/z=430.2 (M⁺).

PREPARATION 7

[0266]

[0267] A solution of compound 6 (0.696 g, 1.617 mmol) in 10 mL of EtOHand 5 mL of THF was hydrogenated at atmospheric pressure at roomtemperature over 70 mg of 10% Pd/C. After 20 h, the mixture was filteredand evaporated to give a solid. Flash chromatography through 50 g ofsilica gel (5:2:2:1 hexane/CH₂Cl₂/EtOAc/MeOH+0.05% HOAc) yielded 0.540 gof the title compound as a white solid.

[0268]¹H-NMR (500 Mz, CDCl₃): δ 1.45 (s, 9H), 2.43 (dd, J=16.6, 5.1 Hz,1H), 2.62 (dd, J=16.8, 8.6 Hz, 1H), 2.84 (dd, J=15.5, 10.4 Hz, 1H),3.1-3.2 (m, 2H), 7.25-7.60(m, 9H). MS (EI): m/z=340.2 (M⁺).

PREPARATION 8

[0269]

[0270] STEP A:

[0271] A stirred solution of compound 3 from Preparation 3 (1.011 g,3.825 mmol) in 15.5 mL of THF was cooled to −70° C. and a 1.0M solutionof LiN(TMS)₂ in hexane (8.42 mL, 8.42 mmol) was added dropwise. After 1h, a freshly prepared 1.14M solution of p-iodobenzyl iodide in THF (6.0mL, 6.84 mmol) was added dropwise. The solution was stirred at −70° C.for 30 min and was then allowed to warm gradually to 10° C. during 90min. The reaction was hydrolyzed by the addition of sat. aqueous NH₄Cland most of the THF was removed by rotary evaporation. The residue waspartitioned between ethyl acetate and sat. aqueous NH₄Cl and the organicphase was washed with water and brine. The organic layer was dried overNa₂SO₄ and evaporated in vacuo to give a solid. Flash chromatographythrough 450 g of silica gel (98:2 CH₂Cl₂/MeOH+0.1% HOAc) yielded 1.78 gof compound 8 as a ˜6:1 mixture of (S,S:R,S) diastereomers (major isomerdepicted).

[0272]¹H-NMR (500 Mz, CDCl₃): 1.32 (s, 9H), 2.74-3.05 (m, 6H), 6.89 (d,J=8.2 Hz, 2H), 7.12-7.28 (m, 5H), 7.56 (d, J=8.4Hz, 2H). MS (EI):m/z=480.4 (M⁺).

[0273] STEP B:

[0274] To a solution of compound 8 (182.0 mg, 0.3789 mmol) in 0.6 mL ofCH₂Cl₂ was added neat trifluoroacetic acid (0.2 mL). The solution wasstirred at room temperature for 4 h, and was then evaporated in vacuo togive an oil. Separation by reverse phase medium pressure chromatographyon RP-18 (40:60 MeCN/0.1% aqueous TFA) gave after lyophilization 103.7mg of the title compound as a white solid.

[0275]¹H-NMR (500 Mz, CD₃OD): 2.84-2.91 (m, 2H), 2.96-3.06 (m, 4H), 6.89(d, J=8.2 Hz, 2H), 7.11-7.25 (m, 5H), 7.55 (d, J=8.2 Hz, 2H). MS (EI):m/z=424.2 (M⁺).

PREPARATION 9

[0276]

[0277] STEP A:

[0278] A stirred solution of compound 7 (26.2 mg, 0.0770 mmol) in 0.7 mLof THF was cooled to −70° C. and a freshly prepared 1.0M solution ofLiN(i-Pr)₂ in THF (0.17 mL, 0.17 mmol) was added dropwise. After 1 h,neat benzyl bromide (0.015 mL, 0.12 mmol) was added dropwise. Thesolution was stirred at −70° C. for 20 min and was then allowed to warmgradually to 10° C. during 90 min. The reaction was hydrolyzed by theaddition of sat. aqueous NH₄Cl. The reaction mixture was partitionedbetween ethyl acetate and sat. aqueous NH₄Cl and the organic phase waswashed with water and brine. The organic layer was dried over Na₂SO₄ andevaporated in vacuo to give an oil. Purification by preparative layerchromatography on silica gel (93:7 CH₂Cl₂/MeOH+0.1% HOAc) yielded 28 mgof compound 10 as an ˜8:1 mixture of (S,S:R,S) diastereomers (majorisomer depicted).

[0279]¹H-NMR (500 Mz, CDCl₃): 1.33 (s, 9H, isomer B, minor), 1.37 (s,9H, isomer A, major), 2.85-3.20 (m, 6H, isomers A & B), 7.10-7.65 (m,14H, isomers A & B). MS (EI): m/z=430.3 (M⁺).

[0280] STEP B:

[0281] To a solution of compound 10 from Step A (10.3 mg, 0.0239 mmol)in 0.3 mL of CH₂Cl₂ was added neat trifluoroacetic acid (0.1 mL). Thesolution was stirred at room temperature for 4 h, and was thenevaporated in vacuo to give an oil. Separation by reverse phase mediumpressure liquid chromatography on RP-18 (45:55 MeCN/0.1% aqueous TFA)gave after lyophilization 5.0 mg of compound 11 and 0.7 mg of compound12 as white solids.

[0282] Compound 11:

[0283]¹H-NMR (500 Mz, CD₃OD): 2.90-2.97 (m, 2H), 3.0-3.1 (m, 4H), 7.14(d, J=7.1 Hz, 3H), 7.15-7.25 (m, 5H), 7.41 (t, J=7.7 Hz, 2H), 7.52 (d,J=8.0 Hz, 2H), 7.58 (d, J=7.8Hz, 2H). MS (EI): m/z=374.2 (M⁺).

[0284] Compound 12:

[0285]¹H-NMR (500 Mz, CD₃OD): 2.85-3.00 (m, 6H), 7.19 (d, J=6.8 Hz, 3H),7.24-7.32 (m, 5H), 7.30 (t, J=7.4 Hz, 1H), 7.41 (dd, J=7.5, 8.0 Hz, 2H),7.49 (d, J=8.0 Hz, 2H), 7.58 (d, J=7.6 Hz, 2H). MS (EI): m/z=374.2 (M⁺).

PREPARATION 10

[0286]

[0287] STEP A:

[0288] A stirred solution of compound 8 (830.1 mg, 1.728 mmol) andp-methoxybenzyl alcohol (0.54 ml, 4.33 mmol) in 14 mL of CH₂Cl₂ wascooled to 0° C., and a 1.0M solution of N,N-dimethylaminopyridine inCH₂Cl₂ (0.259 ml, 0.259 mmol) was added, followed by neat1,3-diisopropylcarbodiimide (0.541 ml, 3.46 mmol). After 1 hr, thecooling bath was removed. The reaction mixture was stirred an additional180 min, and was then hydrolyzed by the addition of sat. aqueous NH₄Cl.The reaction mixture was partitioned between ethyl acetate and sat.aqueoues NH₄Cl and the organic phase was washed with water and brine.The organic layer was dried over Na₂SO₄ and evaporated in vacuo to givea semi-solid. This crude material was triturated with 10 ml CH₂Cl₂ andfiltered through a sintered-glass funnel. Evaporation of the filtrate invacuo gave an oil. Flash chromatography through 160 g of silica gel(73:20:7 hexane/CH₂Cl₂/EtOAc) yielded 921.6 mg of compound 13 as a whitesolid.

[0289]¹H-NMR (500 Mz, CDCl₃): 1.37 (s, 9H), 2.8-3.1 (m, 6H), 3.83 (s,3H), 4.98 (dd, J =43.5, 11.9 Hz, 2H), 6.76 (d, J=8.0 Hz, 2H), 6.86 (d,J=8.4 Hz, 2H), 7.07 (d, J=6.6 Hz, 2H), 7.16-7.27 (m, 5H), 7.53 (d, J=8.1Hz, 2H). MS (ESI): m/z=623.2 (M+Na⁺).

[0290] STEP B:

[0291] To a stirred solution of compound 13 (76.6 mg, 0.1276 mmol) andtetrakis(triphenylphosphine)palladium(0) (7.4 mg, 0.0064 mmol) in 1.1 mlDME was added a solution of 4-methoxybenzeneboronic acid (29.1 mg, 0.192mmol) in 0.2 ml DME. After 10 min, a 2.0M solution of Na₂CO₃ in H₂O(0.130 ml, 0.260 mmol) was added and the reaction mixture was heated to100° C. for 3.5 hr. The reaction mixture was allowed to cool to RT andthen hydrolyzed by the addition of sat. aqueous NH₄Cl. The reactionmixture was partitioned between ethyl acetate and sat. aqueous NH₄Cl andthe organic phase was washed with sat. aqueous NaS₂O₃, water, and brine.The organic layer was dried over Na₂SO₄ and evaporated in vacuo to givean oil. Flash chromatography through 18 g of silica gel (73:20:7hexane/CH₂Cl₂/EtOAc) yielded 37.1 mg of compound 14 as a white solid.

[0292]¹H-NMR (500 Mz, CDCl₃): 1.36 (s, 9H), 2.90-3.07 (m, 6H), 3.82 (s,3H), 3.87 (s, 3H), 4.98 (dd, J=34.8, 11.9 Hz, 2H), 6.86 (d, J=8.7 Hz,2H), 6.99 (d, J=8.7 Hz, 2H), 7.09 (d, J=7.8 Hz, 4H), 7.16-7.26 (m, 5H),7.42 (d, J=8.0 Hz, 2H), 7.52 (d, J=8.5 Hz, 2H). MS (ESI): m/z=603.3(M+Na⁺).

[0293] STEP C:

[0294] To a solution of compound 14 (37.1 mg, 0.064 mmol) in 0.6 mL ofCH₂Cl₂ was added neat trifluoroacetic acid (0.2 mL). The solution wasstirred at room temperature for 4 h, and was then evaporated in vacuo togive an oil. Separation by reverse phase medium pressure chromatographyon RP-18 (45:55 MeCN/0.1% aqueous TFA) gave after lyophilization 9.3 mgof the title compound as a white solid.

[0295]¹H-NMR (500 Mz, CD₃OD): 2.89-2.96 (m, 2H), 3.01-3.07 (m, 4H), 3.81(s, 3H), 6.97 (d, J=8.9Hz, 2H), 7.11-7.25 (m, 7H), 7.43 (d, J=8.2Hz,2H), 7.51 (d, J=8.7 Hz, 2H). MS (ESI): m/z=427.1 (M+Na⁺).

EXAMPLE 1

[0296] Compound 20

[0297] STEP A:

[0298] A stirred solution of compound 3 (0.253 g, 0.957 mmol) in 7 mL ofTHF was cooled to −78° C. and a 1.0M solution of LiN(TMS)₂ in hexanes(2.2 mL, 2.2 mmol) was added dropwise during 5 min. After 65 min, asolution of 4-(iodomethyl)-4′-(t-butyldimethylsilyloxymethyl)biphenyl(0.630 g, 1.44 mmol) in 1 mL of THF was added dropwise during 5 min. Thesolution was stirred at −78° C. for 35 min and was then allowed to warmgradually to −20° C. during 2 h, at which point the reaction was judgedto be complete by HPLC analysis. The reaction was hydrolyzed by theaddition of sat. aqueous NH₄Cl. The reaction mixture was thenpartitioned between ethyl acetate and sat. aqueous NH₄Cl and the organicphase was washed with water and brine. The organic layer was dried overNa₂SO₄ and evaporated in vacuo to give a solid. Flash chromatography onsilica gel (95:3:2 CH₂Cl₂/EtOAc/MeOH) yielded 0.479 g of compound 16 asan ˜19:1 mixture of (S,S:R,S) diastereomers (major isomer depicted).

[0299]¹H-NMR (500 Mz, CDCl₃): δ 0.15 (s, 6H), 0.99 (s, 9H), 1.39 (s,9H), 2.85-2.95 (m, 1H), 2.95-3.20 (m, 5H),4.81 (s, 2H), 7.13 (d, J=6.9Hz, 2H), 7.17 (d, J=8.1 Hz, 2H), 7.20-7.35 (m, 3H), 7.42 (d, J=8.0 Hz,2H), 7.49 (d, J=8.0 Hz, 2H), 7.57 (d, J =8.0 Hz, 2H). MS (EI): m/z=574.3(M⁺).

[0300] STEP B:

[0301] A stirred solution of compound 16 (0.479 g, 0.835 mmol) andp-methoxybenzyl alcohol (0.260 mL, 2.08 mmol) in 7.5 mL of CH₂Cl₂ wascooled to 0° C. and a 1.0 M solution of N,N-dimethylaminopyridine inCH₂Cl₂ (0.125 mL, 0.125 mmol) was added followed by neat1,3-diisopropylcarbodiimide (0.260 mL, 1.66 mmol). After 40 min, thecooling bath was removed. The reaction mixture was stirred for anadditional 140 min, and was then hydrolyzed by the addition of sat.aqueous NH₄Cl. The reaction mixture was partitioned between ethylacetate and sat. aqueous NH₄Cl and the organic phase was washed withwater and brine. The organic layer was dried over Na₂SO₄ and evaporatedin vacuo to give a semi-solid. Flash chromatography on silica gel(75:21.5:3.5 to 75:20:5 hexane/CH₂Cl₂/EtOAc) yielded 0.486 g of compound17 as an oil. The diastereomeric ratio was>100:1 (S,S): (R,S).

[0302]¹H-NMR (500 Mz, CDCl₃): δ 0.16 (s, 6H), 0.99 (s, 9H), 1.37 (s,9H), 2.90-3.10 (m, 6H), 3.83 (s, 3H), 4.81 (s, 2H), 4.96 (d, J=11.9 Hz,1H), 5.04 (d, J=11.9 Hz, 1H), 6.87 (d, J=8.2 Hz, 2H), 7.07-7.15 (m, 4H),7.18-7.28 (m, 5H), 7.41 (d, J=7.8 Hz, 2H), 7.47 (d, J=7.7 Hz, 2H), 7.56(d, J=8.0 Hz, 2H). MS (ESI): m/z=712.6 (M+NH₄ ⁺).

[0303] STEP C:

[0304] A stirred solution of compound 17 (0.482 g, 0.694 mmol) in 4.5 mLof THF was cooled to 0° C. and neat acetic acid (0.120 mL, 2.1 mmol) wasadded followed by a 1.0 M solution of tetrabutylammonium fluoride in THF(2.1 mL, 2.1 mmol). After 50 min, the cooling bath was removed. Afterstirring at room temperature for 22 h, the reaction was judged to becomplete by TLC on silica gel. The reaction mixture was partitionedbetween ethyl acetate and sat. aqueous NH₄Cl and the organic phase waswashed with water and brine. The organic layer was dried over Na₂SO₄ andevaporated in vacuo to give an oil. Flash chromatography on silica gel(4:3:3 hexane/CH₂Cl₂/EtOAc) yielded 0.389 g of compound 18 as an oil.

[0305]¹H-NMR (500 Mz, CDCl₃): δ 1.37 (s, 9H), 2.90-3.10 (m, 6H), 3.82(s, 3H), 4.77 (s, 2H), 4.96 (d, J=11.9 Hz, 1H), 5.03 (d, J=11.9 Hz, 1H),6.87 (d, J=8.5 Hz, 2H), 7.05-7.30 (m, 9H), 7.4-7.5 (m, 4H), 7.59 (d,J=8.0 Hz, 2H). MS (EST): m/z=598.5 (M+NH₄ ⁺).

[0306] STEP D:

[0307] A stirred solution of compound 18 (20.0 mg, 0.0344 mmol) in 0.35mL of CH₂Cl₂ was cooled to −70° C. and neat 1-methylimidazole (0.0090mL, 0.11 mmol) was added followed by trifluoromethanesulfonic anhydride(0.0090 mL, 0.053 mmol). The temperature was allowed to gradually riseand after 45 min was −15° C. TLC on silica gel showed some remainingstarting material, so additional 1-methylimidazole (0.0050 mL, 0.063mmol) was added. After 25 min more, the temperature was 5° C. and TLCshowed no starting material. The reaction mixture was partitionedbetween ethyl acetate and water and the organic phase was washed oncewith water. The organic layer was dried over Na₂SO₄ and evaporated invacuo to give an oil. Preparative layer chromatography on silica gel(1:1 MeCN/CH₂Cl₂) yielded 19.0 mg of compound 19 as an oil.

[0308]¹H-NMR (500 Mz, CDCl₃): δ 1.36 (s, 9H), 2.88-3.08 (m, 6H), 3.81(s, 3H), 3.94 (s, 3H), 4.95 (d, J=12 Hz, 1H), 5.02 (d, J=12 Hz, 1H),5.39 (s, 2H), 6.86 (d, J=8.6 Hz, 2H), 7.05-7.35 (m, 11H), 7.43 (d, J=8.3Hz, 2H), 7.48 (d, J=8.0 Hz, 2H), 7.60 (d, J =8.3 Hz, 2H), 9.23 (s, 1H).MS (ESI): m/z=645.5 (M⁺).

[0309] STEP E:

[0310] To a solution of compound 19 (18.5 mg, 0.0233 mmol) in 0.3 mL ofCH₂Cl₂ was added neat trifluoroacetic acid (0.1 mL). The solution wasstirred at room temperature for 110 min, and was then evaporated invacuo to give a solid. Purification by reverse phase medium pressureliquid chromatography on RP-18 (40:60 MeCN/0.1% aqueous TFA) gave afterlyophilization 10.4 mg of compound 20 as a white solid.

[0311]¹H-NMR (500 Mz, CD₃OD): δ 2.88-2.98 (m, 2H), 3.0-3.1 (m, 4H), 3.93(s, 3H), 5.44 (s, 2H), 7.10-7.25 (m, 7H), 7.45-7.55 (m, 4H), 7.59 (s,1H), 7.65 (s, 1H), 7.68 (d, J=8.2 Hz, 2H), 8.99 (s, 1H). MS (ESI):m/z=469.4 (M⁺).

EXAMPLE 2

[0312] Compound 23

[0313] STEP A:

[0314] A stirred solution of compound 18 (0.262 g, 0.451 mmol) in 4.5 mLof CH₂Cl₂ was cooled to −60° C. and triethylamine (0.107 mL, 0.768 mmol)was added followed by neat methanesulfonyl chloride (0.0490 mL, 0.633mmol). The temperature was allowed to gradually rise and after 45 minwas −25° C. The reaction was hydrolyzed by the addition of sat. aqueousNH₄Cl. The reaction mixture was partitioned between ethyl acetate andsat. aqueous NH₄Cl and the organic phase was washed with water andbrine. The organic layer was dried over Na₂SO₄ and evaporated in vacuoto give a solid.

[0315] The crude mesylate was dissolved in acetone and cooled to 0° C.Solid sodium iodide was added (0.135 g, 0.901 mmol) and the mixture wasstirred in the dark. After 30 min, the cooling bath was removed. Afterstirring for 2 h more, the reaction mixture was partitioned betweenethyl acetate and water. The organic phase was washed with 51% aqueousNa₂S₂O₃, water and brine. The organic lawyer was dried over Na₂SO₄ andevaporated in vacuo to give 313 mg of compound 21 as a solid.

[0316]¹H-NMR (500 Mz, CDCl₃): δ 1.36 (s, 9H), 2.9-3.1 (m, 6H), 3.82 (s,3H), 4.54 (s, 2H), 4.96 (d, J=11.9 Hz, 1H), 5.03 (d, J=11.9 Hz, 1H),6.87 (d, J=8.5 Hz, 2H), 7.06-7.14 (m, 4H), 7.17-7.28 (m, 5H), 7.42-7.50(m, 4H), 7.51 (d, J=8.2Hz, 2H). MS (ESI): m/z=708.1 (M+NH₄ ⁺).

[0317] STEP B:

[0318] To a stirred solution of compound 21 (0.0212 g, 0.0307 mmol) and1-(aminocarbonylmethyl-4-aza-1-azoniabicyclo(2.2.2)octanetrifluoromethanesulfonate (0.011 g, 0.034 mmol) in 0.35 mL ofacetonitrile and 0.1 mL of THF was added a solution of silvertrifluoromethanesulfonate in acetonitrile (0.845 M, 0.036 mL, 0.030mmol). A precipitate formed immediately. The mixture was stirred in thedark for 55 min and was then filtered and evaporated in vacuo to give 37mg of compound 22 as a solid.

[0319]¹H-NMR (500 Mz, d₆-acetone): δ 1.32 (s, 9H), 2.9-3.1 (m, 6H), 3.80(s, 3H), 4.38-4.48 (m, 6H), 4.50-4.60 (m, 6H), 4.65 (s, 2H), 4.93 (d,J=12 Hz, 1H), 5.02 (d, J=12 Hz, 1H), 5.14 (s, 2H), 6.90 (d, J=8.7 Hz,2H), 7.16 (d, J=6.9 Hz, 2H), 7.2-7.3 (m, 8H), 7.63 (d, J=8.0 Hz, 2H),7.69 (bs, 1H), 7.80 (d, J=8.2 Hz, 2H), 7.86 (d, J=8.2 Hz, 2H). MS (ESI):m/z=732.5 (M⁺²−H⁺).

[0320] STEP C:

[0321] A solution of compound 22 (37 mg) in 0.3 mL of CH₂Cl₂ and 0.1 mLof trifluoroacetic acid was stirred at room temperature for 150 min, andwas then evaporated in vacuo to give a film. Purification by reversephase medium pressure liquid chromatography on RP-18 (25:75 MeCN/0.1%aqueous TFA) gave after lyophilization 21.8 mg of compound 23 as a whitesolid.

[0322]¹H-NMR (500 Mz, CD₃OD): δ 2.88-2.98 (m, 2H), 3.00-3.15 (m, 4H),3.18-4.08 (m, 6H), 4.2-4.3 (m, 6H), 4.36 (s, 2H), 4.82 (s, 2H), 7.14 (d,J=7.1 Hz, 2H), 7.15-7.30 (m, 5H), 7.56 (d, J=8.0 Hz, 2H), 7.63 (d, J=8.2Hz, 2H), 7.82 (d, J=8.3 Hz, 2H). MS (ESI): m/z=556.4 (M⁺²−H⁺).

EXAMPLE 3

[0323] Compound 25

[0324] STEP A:

[0325] To a stirred solution of compound 21 (0.194 g, 0.281 mmol) in0.75 mL of THF was added a solution of1-(1-azidoprop-3-yl)-4-aza-1-azoniabicyclo(2.2.2)octanetrifluoromethanesulfonate (0.107 g, 0.310 mmol) in 2.25 mL ofacetonitrile. To the resulting solution was added a solution of silvertrifluoromethanesulfonate in acetonitrile (0.845 M, 0.332 mL, 0.281mmol). A precipitate formed immediately. The mixture was stirred in thedark for 45 min and was then filtered and evaporated in vacuo to give312 mg of compound 24 as a solid. ¹H-NMR (500 Mz, d₆-acetone): δ 1.32(s, 9H), 2.25-2.35 (m, 2H), 2.90-3.15 (m, 6H), 3.55-3.65 (m, 2H), 3.80(s, 3H), 3.93-4.00 (m, 2H), 4.37 (s, 12H), 4.93 (d, J=12 Hz, 1H), 5.02(d, J=12 Hz, 1H), 5.14 (s, 2H), 6.90 (d, J=8.4 Hz, 2H), 7.16 (d J=6.6Hz, 2H), 7.2-7.3 (m, 7H), 7.63 (d, J=8.2 Hz, 2H), 7.80 (d, J 8.5 Hz,2H), 7.85 (d, J=8.2 Hz, 2H). MS (ESI): m/z=872.1 (M⁺²+CF₃CO₂—).

[0326] STEP B:

[0327] A solution of compound 24 (312 mg) in 2.1 mL of CH₂Cl₂ and 0.7 mLof trifluoroacetic acid was stirred at room temperature for 125 min, andwas then evaporated in vacuo. The resulting dry film was dissolved in 3mL of THF, 1 mL of MeOH and 1 mL of water and hydrogenated atatmospheric pressure over 55 mg of 5% rhodium on carbon. After 3 h, themixture was filtered through a pad of Celite®. The tan filtrate wasconcentrated by rotary evaporation until it became hazy, and was thenfrozen and lyophilized to give a brown solid. Purification by reversephase medium pressure liquid chromatography on RP-18 (20:80 MeCN/0.1%aqueous TFA) gave after lyophilization 164 mg of a white solid. Aportion of this solid (151.4 mg) was dissolved in 2 mL of methanol andeluted with methanol through a 12 g column of Bio-Rad® AG-2-X8 chlorideform resin (˜3 meq/g). Evaporation of the collected fractions gave acolorless oil which was lyophilized from water/MeCN to give 109 mg ofcompound 25 as a white solid.

[0328]¹H-NMR (500 Mz, CD₃OD): δ 2.18-2.30 (m, 2H), 2.9-3.0 (m, 2H),3.00-3.15 (m, 6H), 3.7-3.8 (m, 2H), 4.08 (s, 12H), 4.89 (s, 2H), 7.14(d, J=7.0 Hz, 2H), 7.15-7.30 (m, 5H), 7.56 (d, J=8.0 Hz, 2H), 7.67 (d,J=8.0 Hz, 2H), 7.82 (d, J=8.0Hz, 2H). MS (ESI): m/z=556.4 (M⁺³−2H⁺).

EXAMPLE 4

[0329] Compound 27

[0330] STEP A:

[0331] To a stirred solution of compound 21 (41.2 mg, 0.0597 mmol) and[3-(dimethylamino)propyl] [3-[(1,1-dimethylethoxy)carbonylamino]propyl]carbamic acid, 1,1-dimethylethyl ester (24.7 mg, 0.0687mmol) in 1.2 mL of acetonitrile was added a solution of silvertrifluoromethanesulfonate in acetonitrile (0.845 M, 0.066 mL, 0.0564mmol). A precipitate formed immediately. The mixture was stirred in thedark for 85 min and was then filtered and evaporated in vacuo to give66.7 mg of compound 26 as a solid.

[0332] 1H-NMR (500 Mz, d₆-acetone): δ 1.33 (s, 9H), 1.40 (s, 9H), 1.45(s, 9H), 1.74 (bs, 2H), 2.37 (bs, 2H), 2.9-3.2 (m, 8H), 3.26-3.33 (m,2H), 3.35 (s, 6H), 3.42 (bs, 2H), 3.61 (bs, 2H), 3.80 (s, 3H), 4.85 (s,2H), 4.97 (dd, J=43.5, 11.9 Hz, 2H), 5.99 (bs, 1H), 6.91 (d, J=8.0 Hz,2H), 7.16 (d, J=7.3 Hz, 2H), 7.19-7.30 (m, 7H), 7.63 (d, J=7.5 Hz, 2H),7.8 (dd, J=37.1, 7.9 Hz, 4H). MS (ESI): m/z=922.5 (M⁺).

[0333] STEP B:

[0334] To a solution of compound 26 (66.7 mg) in 0.6 mL of CH₂Cl₂ wasadded neat trifluoroacetic acid (0.2 mL). The solution was stirred atroom temperature for 5.5 h, and was then evaporated in vacuo to give anoil. Purification by reverse phase medium pressure chromatography onRP-18 (30:70 MeCN/0.1% aqueous TFA) gave after lyophilization 36 mg of awhite solid. A portion of this solid (31.5 mg) was dissolved in 2 mL ofmethanol and eluted with methanol through a 3 g column of Bio-Rad®AG-2-X8 chloride form resin (˜3 meq/g). Evaporation of the collectedfractions gave a colorless oil which was lyophilized from water/MeCN togive 22.2 mg of compound 27 as a white solid.

[0335] 1H-NMR (500 Mz, CD₃OD): δ 2.07-2.16 (m, 2H), 2.30-2.40 (m, 2H),2.90-2.97 (m, 2H), 3.03-3.22 (m, 16H), 3.43-3.50 (m, 2H), 4.60 (s, 2H),7.12-7.28 (m, 7H), 7.56 (d, J=8.2 Hz, 2H), 7.63 (d, J=8.2 Hz, 2H), 7.78(d, J=8.2 Hz, 2H). MS (ESI): m/z=546.4 (M⁺³−2H⁺).

EXAMPLES 5-29

[0336] Employing the procedures described herein, additional compoundsof the present invention were prepared. These are described in Tables3-7, which additionally include characterizing data. TABLE 3

Example No. Q^(⊕) Y^(⊖) m/z 5

3 Cl^(⊖) 542.5(M⁺³ − 2H⁺); ESI 6

3 Cl^(⊖) 660.4(M⁺³ − H⁺ + CF₃CO₂ ⁻); ESI 7

3 Cl^(⊖) 532.3(M⁺³ − 2H⁺); ESI 8

4 Cl^(⊖) 613.4(M⁺⁴ − 3H⁺); ESI 9

4 Cl^(⊖) 10

3 Cl^(⊖) 649.5(M⁺² − H⁺); ESI 11

3 Cl^(⊖) 712.3(M⁺³ − H⁺ + CF₃CO₂—); ESI 12

3 Cl^(⊖) 765.5(M⁺³ − H⁺ + CF₃CO₂ ⁻); ESI 13

3 Cl^(⊖) 599.3(M⁺³ − 2H⁺); ESI 14

3 Cl^(⊖) 793.5(M⁺³ − H⁺ + CF₃CO₂ ⁻); ESI

[0337] TABLE 4

Example No. Q^(⊕) Y^(⊖) m/z 15

CF₃CO₂ ^(⊖) 393.3(M⁺); EI 16

CF₃CO₂ ^(⊖) 328.1(M⁺); EI 17

2 Cl^(⊖) 437.8(M⁺); ESI 18

2 CF₃CO₂ ^(⊖) 480.6(M⁺² − H⁺); ESI 19

3 CF₃CO₂ ^(⊖) 480.5(M⁺³ − 2H⁺); ESI 20

3 CF₃CO₂ ^(⊖) 523.4(M⁺³ − 2H⁺); ESI

[0338] TABLE 5

Example No. Q^(⊕) Y^(⊖) m/z 21

Cl^(⊖) 421.5(M⁺); ESI 22

2 Cl^(⊖) 508.6(M⁺² − H⁺); ESI 23

3 Cl^(⊖) 508.3(M⁺³ − 2H⁺); ESI 24

3 Cl^(⊖) 551.3(M⁺³ − 2H⁺); ESI

[0339] TABLE 6

Example No. Q^(⊕) Y^(⊖) m/z 25

Cl^(⊖) 387.5(M⁺); ESI 26

2 Cl^(⊖) 474.7(M⁺² − H⁺); ESI 27

3 Cl^(⊖) 474.4(M⁺³ − 2H⁺); ESI 28

3 Cl^(⊖) 517.4(M⁺³ − 2H⁺); ESI

[0340] TABLE 7

Example No. Q^(⊕) Y^(⊖) m/z 29

Cl^(⊖) 469.3(M⁺); ESI

EXAMPLES 30-104

[0341] Additional compounds of the present invention can be preparedemploying the procedures described herein. These are described in Tables8-18. TABLE 8

Example No. Q^(⊕) Y^(⊖) 30

3 Cl^(⊖) 31

3 CH₃CO₂ ^(⊖) 32

3 Cl^(⊖) 33

3 Cl^(⊖) 34

3 Cl^(⊖) 35

3 Cl^(⊖) 36

4 Cl^(⊖) 37

4 CH₃CO₂ ^(⊖) 38

4 Cl^(⊖) 39

4 Cl^(⊖) 40

4 Cl^(⊖) 41

4 Cl^(⊖) 42

Cl^(⊖) 43

Cl^(⊖) 44

3 Cl^(⊖) 45

3 Cl^(⊖) 46

4 Cl^(⊖) 47

3 Cl^(⊖)

[0342] TABLE 9

Example No. Q^(⊕) Y^(⊖) 48

Cl^(⊖) 49

2 Cl^(⊖) 50

3 Cl^(⊖) 51

3 Cl^(⊖) 52

3 Cl^(⊖) 53

3 Cl^(⊖)

[0343] TABLE 10

Example No. Q^(⊕) Y^(⊖) 54

Cl^(⊖) 55

2 Cl^(⊖) 56

3 Cl^(⊖) 57

3 Cl^(⊖) 58

3 Cl^(⊖) 59

4 Cl^(⊖)

[0344] TABLE 11

Example No. Q^(⊕) Y^(⊖) 60

Cl^(⊖) 61

2 Cl^(⊖) 62

3 Cl^(⊖) 63

3 Cl^(⊖) 64

3 Cl^(⊖) 65

4 Cl^(⊖)

[0345] TABLE 12

Example No. Q^(⊕) Y^(⊖) 66

Cl^(⊖) 67

2 Cl^(⊖) 68

3 Cl^(⊖) 69

3 Cl^(⊖) 70

3 Cl^(⊖) 71

4 Cl^(⊖)

[0346] TABLE 13

Example No. Q^(⊕) Y^(⊖) 72

Cl^(⊖) 73

2 Cl^(⊖) 74

3 Cl^(⊖) 75

3 Cl^(⊖) 76

3 Cl^(⊖) 77

4 Cl^(⊖)

[0347] TABLE 14

Example No. Q^(⊕) Y^(⊖) 78

Cl^(⊖) 79

2 Cl^(⊖) 80

3 Cl^(⊖) 81

3 Cl^(⊖) 82

3 Cl^(⊖) 83

4 Cl^(⊖)

[0348] TABLE 15

Example No. Q^(⊕) Y^(⊖) 84

Cl^(⊖) 85

2 Cl^(⊖) 86

3 Cl^(⊖) 87

3 Cl^(⊖) 88

3 Cl^(⊖) 89

4 Cl^(⊖)

[0349] TABLE 16

Example No. Q^(⊕) Y^(⊖) 90

Cl^(⊖) 91

2 Cl^(⊖) 92

3 Cl^(⊖) 93

3 Cl^(⊖) 94

3 Cl^(⊖) 95

4 Cl^(⊖)

[0350] TABLE 17

Example No. Q^(⊕) Y^(⊖) 96

Cl^(⊖) 97

2 Cl^(⊖) 98

3 Cl^(⊖) 99

3 Cl^(⊖) 100

3 Cl^(⊖) 101

4 Cl^(⊖)

[0351] TABLE 18 Example No. 102

103

104

BIOLOGICAL ACTIVITY

[0352] IMP-1 metallo-β-lactamase lacking the N-terminal 18 hydrophobicamino acids which encode the putative periplasmic signal sequence (EMBLaccess code PACATAAC6) was PCR amplified from plasmid DNA prepared froma carbapenem-resistant strain of Pseudomonas aeruginosa (CL5673). ThePCR product was cloned into pET30a+ (Novegen) and expressed in E. coliBL21(DE3) after induction with 0.5 mM IPTG for 20 hours at roomtemperature in minimal media supplemented with casamino acids and 348 μMZnSO₄. Soluble IMP-1 was purified from cell extracts by SP-Sepharose(Pharmacia) ion exchange and Superdex 75 (Pharmacia) size-exclusionchromatography. Soluble CcrA metallo-β-lactamase was cloned from animipenem resistant clinical isolate of Bacteroides fragilis and wasexpressed and purified as described by Toney et al. [Protein Expr.Purif. 9 355 (1997)].

[0353] The IC₅₀ of succinate derivatives was determined following a 15minute incubation at 37° C. with IMP-1 (0.75 nM in 50 mM MOPS, pH 7) orCcrA (4 nM in 50 mM Mops pH 7). Using initial velocity as a measure ofactivity, inhibition was monitored spectrophotometrically at 490 nm in aMolecular Devices SPECTRAmax™ 250 96-well plate reader employingnitrocefin as the reporter substrate at approximately K_(m)concentration (60 μM).

[0354] A laboratory strain of E. coli engineered to express IMP-1 wasused to evaluate the ability of succinate derivatives to reversemetallo-β-lactamase-mediated carbapenem resistance in bacteria. NativeIMP-1, which included the N-terminal periplasmic signal sequence, wasPCR amplified from CNA isolated from a carbapenem resistant P.aeruginosa clinical isolate, CL56673, and cloned into the pET30a vector.The basal (uninduced) level of IMP-1 expressed when pET30a-IMP-1 wasintroduced into E. coli BL21(DE3) resulted in 4-, 64- or 500-foldreduced sensitivity to impenem, meropenem or(1S,5R,6S)-1-methyl-2-{7[-4-(aminocarbonylmethyl)-1,4-diazoniabicyclo(2.2.2)octan-1-yl]methyl-fluoren-9-on-3-yl}-6-(1R-hydroxyethyl)-carbapen-2-em-3-carboxylatechloride (a carbapenem synthesized at Merck Research Laboratories)respectively. For example, the minimum inhibitory concentration (MIC) of(1S,5R,6S)-1-methyl-2-{7-[4-(aminocarbonylmethyl)-1,4-diazoniabicyclo(2.2.2)octan-1-yl]methyl-fluoren-9-on-3-yl}-6-(1R-hydroxyethyl)-carbapen-2-em-3-carboxylatechloride, was typically increased from 0.06-0.12 μg/ml to 16-32 μg/ml bythe expression of IMP-1. To evaluate IMP-1 inhibitors, an overnightculture of E. coli BL2(DE3)/pET30a-IMP-1, grown 35° C. in LB broth(Difco) or Mueller Hinton broth (BBL) supplemented with kanamycin (50μM/ml), was diluted to a final concentration of ˜10⁵ cells/ml in MuellerHinton broth (BBL) containing a subinhibitory concentration (0.25×MIC)of the carbapenem,(1S,5R,6S)-1-methyl-2-{7-[4-(aminocarbonylmethyl)-1,4-diazoniabicyclo(2.2.2)octan-1-yl]methyl-fluoren-9-on-3-yl}-6-(1R-hydroxyethyl)-carbapen-2-em-3-carboxylatechloride. Various concentrations of IMP-1 inhibitor were added to thebacterial growth medium and their capacity to effect a four-fold orgreater increase in sensitivity to the carbapenem was monitored. Thereadout for antibacterial activity showed no visible growth after 20hours incubation at 35° C.

[0355] Representative compounds of Formula I were tested as inhibitorsagainst purified IMP-1 metallo-β-lactamase and found to be active in anIC₅₀ range of from about 0.1 nM to about 1000 nM. The ability ofrepresentative compounds of Formula I to potentiate the activity of thecarbapenem antibiotic(1S,5R,6S)-1-methyl-2-{7-[4-(aminocarbonylmethyl)-1,4-diazoniabicyclo(2.2.2)octan-1yl]-methyl-fluoren-9-on-3-yl}-6-(1R-hydroxyethyl)-carbapen-2-em-3-carboxylatechloride against an IMP-1 producing laboratory strain E. coliBL21(DE3)/pET30a-IMP-1 was tested. Compounds of Formula I in theconcentration range of from about 0.003 μM to about 12.5 μM. were foundto produce 4-fold increase in sensitivity to the carbapenem antibiotic(1S,5R,6S)-1-methyl-2-{7-[4-(aminocarbonylmethyl)-1,4-diazoniabicyclo(2.2.2)octan-1-yl]-methyl-fluoren-9-on-3-yl}-6-(1R-hydroxyethyl)-carbapen-2-em-3-carboxylatechloride in an IMP-1 producing laboratory strain E. Coli.BL21(DE3)/pET30a-IMP-1.

What is claimed is:
 1. A compound represented by formula I:

including pharmaceutically acceptable salts, prodrugs, anhydrides, andsolvates thereof, wherein: M¹ and M² are independently selected from:(a) Hydrogen, (e) Pharmaceutically acceptable cation, (e)Pharmaceutically acceptable esterifying group; and (g) A negativecharge; R¹ and R² are independently selected from the following: (d)Hydrogen, provided that R¹ and R² are not hydrogen at the same time; (e)a C₁ to C₁₆ straight, branched or unsaturated alkyl group substitutedwith 0 to 2 R^(q) groups and substituted with 0 to 3 R_(x) groups andoptionally interrupted by one of the following O, S, SO₂, —C(O)—, (f)—C(O)—NR^(a)—, —CO₂—; (c) a group of the formula:

 wherein —A— represents a single bond, C₁ to C₈ straight, branched orunsaturated alkyl group optionally substituted with 1 to 2 R_(x) groupsand optionally interrupted by one of the following O, S, SO₂, —C(O)—,—C(O)—NR^(a)—, —CO₂—;

represents: (1) a C₆ to C₁₄ aryl group; (2) a C₃ to C₁₀ alicyclic group;(3) a C₃ to C₁₄ heteroaryl group, which contains 1 to 3 heteroatoms, 0to 3 of which heteroatoms are nitrogen and 0 to 1 of which are oxygen orsulfur; (4) a C₃ to C₁₀ heterocyclic group, which contains 1 to 2heteroatoms, 0 to 1 of which heteroatoms are nitrogen, and 0 to 2 ofwhich are oxygen or sulfur; or (d) a group of the formula:

 wherein: —A— is as defined above; A′ is a single bond, O, S, or a C₁ toC₆ straight, branched or unsaturated alkyl group optionally substitutedwith 1-2 R_(x) groups and optionally interrupted by one of the followinggroups O, S, SO₂, —C(O)—, —C(O)—NR^(a)—, —CO₂—;

are independently selected from: (1) a C₆ to C₁₀ aryl group; (2) a C₃ toC₈ alicyclic group; (3) a C₂ to C₉ heteroaryl group, which contains 1 to3 heteroatoms, 0 to 3 of which heteroatoms are nitrogen and 0 to 1 ofwhich are oxygen or sulfur; (4) a C₃ to C₈ heterocyclic group, whichcontains 1 to 2 heteroatoms, 0 to 1 of which heteroatoms are nitrogen,and 0 to 2 of which are oxygen or sulfur; provided that at least oneR^(q) group is present in R¹ or R² and that when more than one R^(q) ispresent the total number of cationic nitrogen atoms does not exceed 8;the total number of cationic nitrogen atoms can be charged balanced byM¹ and/or M² or by M¹ and/or M² in combination with an appropriatenumber of Y⁻; wherein: R^(q) is —E—Q⁺Y⁻; Y⁻ is a pharmaceuticallyacceptable anionic group; E is —(CH₂)_(m)—X—(CH₂)_(n)—; m is 0 to 6; nis 0 to 6 (but when E is attached to an aromatic ring n is 1-6); X is abond, O, S, SO₂, —C(O)—, —C(O)—N(R^(a))—, —C(O)O—, —CH═CH— or —C≡C—,provided that when X is O, S, —C(O)—N(R^(a))— or —C(O)O—, then n is 2 to6 and Q⁺, attached to the (CH₂)_(n) terminus of E is: (1) a cationicgroup selected from the following:

 wherein: R^(u) and R^(v) are independently hydrogen or C₁₋₆ alkyloptionally substituted with 1 to 2 R^(y); R^(w) is hydrogen or C₁₋₆alkyl optionally substituted with 1 to 2 R_(x); R^(u) and R^(v) whenbonded to the same nitrogen atom may together be a C₃₋₆ alkyl radical,which when taken together with the intervening atoms form a ring; TwoR^(u) groups on separate nitrogen atoms may together comprise a C₂₋₅alkyl radical, which when taken together with the intervening atoms forma ring; R^(u), R^(v) and R^(w) when bonded to the same nitrogen atom maytogether form a C₆₋₁₀ tertiary alkyl radical, which with N⁺ forms abicyclic ring; (2) A dicationic group:

 wherein: E¹ is —(CH₂)_(p)—Z—(CH₂)_(r)—; p and r are independently 1 to4; Z is a bond, O, S, SO₂, —C(O)—, —C(O)O—**, —CH═CH—, —C≡C—, or

Provided that when Z is O or S, p is 2 to 4 and r is 2 to 4 and when Zis

or —C(O)O—**, r is 2 to 4; wherein ** denotes the atom which is bondedto the —(CH₂)_(r)— moiety of E¹ above; Q¹ is selected from thefollowing:

Q² is selected from the following:

 R^(u), R^(v) and R^(w) are independently selected and defined as above,And in addition, in the case where two R^(u) groups on separate nitrogenatoms are joined to form a ring as defined above, two R^(v) groups onthe same two separate nitrogen atoms may also comprise a C₁₋₅ alkylradical to form together with the intervening atoms a bicyclic ring; anexample of such is:

(5) A tricationic group selected from the following:

 wherein: Each E¹ is as defined above, but selected independently; EachQ¹ is as defined above, but selected independently; Each Q² is asdefined above, but selected independently; R^(u), R^(v) and R^(w) aredefined as in the definition of Q⁺ item (2) above and selectedindependently; or (6) A tetracationic group selected from the following:

 wherein: Each E¹ is as defined above, but selected independently; EachQ¹ is as defined above, but selected independently; Each Q² is asdefined above, but selected independently; R^(u), R^(v) and R^(w) aredefined as in the definition of Q⁺ item (2) above and selectedindependently; where each R_(x) is independently selected from the groupconsisting of: (f) F, Cl, Br, I, (g) CF₃, (h) OR^(b), (i) CN, (j)—C(O)—R^(c), (f) —S(O₂)—R^(f), (g) —C(O)—OR^(a) (h) —O—C(O)—R^(c), (i)—S—R^(b), (j) —N(R^(a))—C(O)—R^(c),

(q) —N(R^(a))—C(O)—OR^(f), (r) —S(O)—R^(f), (s) —N(R^(a))—S(O₂)—R^(f),(t) NO₂, and (u) C₁ to C₈ straight, branched or unsaturated alkyloptionally substituted with one of the substituents (a) through (t)above; (v) —CH₂-aryl wherein the aryl is optionally substituted with oneof the substituents (a) through (t) above; or two adjacent R_(x) groupson an aromatic ring may consist of the following divalent moiety,—O—CH₂—O—; wherein: R^(a) is H, C₁ to C₆ alkyl optionally substitutedwith R^(y); R^(b) is H, C₁ to C₆ alkyl optionally substituted withR^(y), CH₂-aryl, or aryl, said aryls optionally substituted with 1-2R^(y) groups; R^(c) is H, C₁ to C₆ alkyl optionally substituted withR^(y), CF₃, or aryl, said aryl optionally substituted with 1-2 R^(y)groups; R^(d) and R^(e) are independently hydrogen, C₁ to C₄ alkyloptionally substituted with R^(y), or R^(d) and R^(e) taken together mayrepresent a 3 to 5-membered alkyl radical to form a ring, or R^(d) andR^(e) taken together may represent a 2 to 4-membered alkyl radicalinterrupted by O, S, SO or SO₂ to form a ring; R^(f) is C₁ to C₆ alkyloptionally substituted with R^(y), or aryl, said aryl optionallysubstituted with 1-2 R^(y) groups; and R^(y) is —OH, —OCH₃, OCONH₂,OCOCH₃, CHO, COCH₃, CO₂CH₃, CONH₂, CN, SOCH₃, SO₂CH₃, SO₂NH₂, F, Cl, Br,I or CF₃.
 2. A compound in accordance with claim 1 wherein M¹ and M² areindependently hydrogen or a negative charge and all other variables areas defined above.
 3. A compound in accordance with claim 1 wherein R¹and/or R² represents a C₁ to C₁₆ straight, branched or unsaturated alkylgroup substituted with 0 to 2 R^(q), and substituted with 0 to 3 R_(x)groups, provided that at least one of R¹ or R² contains an R^(q) and allother variables are defined as above.
 4. A compound in accordance toclaim 1 where R¹ and/or R² represents

wherein at least one R^(q) group is present on R¹ or R² and all othervariables are defined as above.
 5. A compound in accordance to claim 1where R¹ and/or R² represents (d)

wherein at least one R^(q) group is present on R¹ or R² and all othervariables are defined as above.
 6. A compound in accordance with claim 1wherein the relative and absolute stereochemistry is:


7. A compound in accordance with claim 6 wherein R¹ and/or R² representsC₄₋₁₂ straight, branched or unsaturated alkyl group optionallysubstituted with 1 to 2 R_(x) and optionally substituted with 1 to 2R^(q) groups provided that at least one of R¹ or R² contains an R^(q)and all other variables are as described above.
 8. A compound inaccordance with claim 6 wherein R¹ and/or R² represents a group of theformula:

wherein A is (CH₂)₁₋₅ and

is phenyl, naphthyl, cyclohexyl or dibenzofuranyl, provided that atleast one of R¹ or R² contains an R^(q) and all other variables are asoriginally defined.
 9. A compound in accordance with claim 6 whereinwhere R¹ or R² represents a group of the formula:

wherein A is (CH₂)₁₋₃, A′ is a single bond, —O— or (CH₂)₁₋₂ and

 independently represent phenyl, thienyl, pyridyl, furanyl orcyclohexyl.
 10. A compound in accordance with claim 6 where one of R¹ orR² is C₄₋₈ straight, branched or unsaturated alkyl optionallysubstituted with 1 to 2 R_(x) or a group of the formula:

where A is (CH₂)₁₋₂ and

is phenyl, cyclopentyl or cyclohexyl and the other of R¹ or R² is: i) aC₇₋₁₂ alkyl group substituted with R^(q), ii) a group of the formula:

 where A is (CH₂)₁₋₂, A′ is a single bond,

is phenyl, thienyl or cyclohexyl and

is phenyl, thienyl or pyridyl, or iii) a group of the formula:

 where A is (CH₂)₁₋₃,

is phenyl or thienyl and R^(q) is —(CH₂)₂₋₆—Q⁺ Y^(Q); and all othervariables are as originally defined.
 11. A compound according to claim 6wherein R¹ is C₅₋₇ alkyl substituted with 0 to 2 R_(x) goups,

R² is C₇₋₁₀ alkyl substituted with 1 R^(q) groups and 0 to 2 R_(x)groups,

and all other variables are as originally defined.
 12. A compoundaccording to claim 11 wherein R¹ is:

R² is:

and all other variables are as originally defined.
 13. A compoundaccording to claim 1 wherein 1 or 2 R^(q) groups are present containinga total number of 2 to 6 cationic nitrogen atoms.
 14. A compoundaccording to claim 1 wherein a single R^(q) substituent is presentcontaining a tricationic or tetracationic Q⁺ group.
 15. A compoundaccording to claim 14 wherein R^(q) is —E—Q⁺Y⁻ wherein E is (CH₂)₀₋₆ or—C(O)—N(R^(a))—(CH₂)₂₋₄— and Q⁺ is a tricationic or tetracationic groupand Y⁻ and R^(a) are as originally defined.
 16. A compound according toclaim 14 wherein the tricationic Q⁺ group is selected from the groupconsisting of:

wherein E¹ is (CH₂)₂₋₄ or —(CH₂)—C(O)—N(R^(a))—(CH₂)₂₋₄— and R^(a), Q¹and Q² are as previously defined.
 17. A compound according to claim 16wherein the tricationic Q⁺ group is selected from:

wherein R^(u), R^(v), and R^(w) are as defined above.
 18. A compoundaccording to claim 14 wherein the tetracationic Q⁺ group is selectedfrom the group consisting of:

wherein E¹ is (CH₂)₂₋₄ or —(CH₂)—C(O)—N(R^(a))—(CH₂)₂₋₄— and R^(a), Q¹,Q², and R^(w) are as defined above.
 19. A compound according to claim 18wherein the tetracationic Q⁺ group is selected from:

wherein R^(a), R^(u), R^(v), and R^(w) are as described above.
 20. Acompound of the structural formula:


21. A compound represented by Tables 3-7: TABLE 3

Example No. Q^(⊕) Y^(⊖) 5

3 Cl^(⊖) 6

3 Cl^(⊖) 7

3 Cl^(⊖) 8

4 Cl^(⊖) 9

4 Cl^(⊖) 10

3 Cl^(⊖) 11

3 Cl^(⊖) 12

3 Cl^(⊖) 13

3 Cl^(⊖) 14

3 Cl^(⊖)

TABLE 4

Example No. Q^(⊕) Y^(⊖) 15

CF₃CO₂ ^(⊖) 16

CF₃CO₂ ^(⊖) 17

2 Cl^(⊖) 18

2 CF₃CO₂ ^(⊖) 19

3 CF₃CO₂ ^(⊖) 20

3 CF₃CO₂ ^(⊖)

TABLE 5

Example No. Q^(⊕) Y^(⊖) 21

Cl^(⊖) 22

2 Cl^(⊖) 23

3 Cl^(⊖) 24

3 Cl^(⊖)

TABLE 6

Example No. Q^(⊕) Y^(⊖) 25

Cl^(⊖) 26

2 Cl^(⊖) 27

3 Cl^(⊖) 28

3 Cl^(⊖)

TABLE 7

Example No. Q^(⊕) Y^(⊖) 29

Cl^(⊖)


22. A compound represented by Tables 8-18: TABLE 8

Example No. Q^(⊕) Y^(⊖) 30

3 Cl^(⊖) 31

3 CH₃CO₂ 32

3 Cl^(⊖) 33

3 Cl^(⊖) 34

3 Cl^(⊖) 35

3 Cl^(⊖) 36

4 Cl^(⊖) 37

4 CH₃CO₂ ^(⊖) 38

4 Cl^(⊖) 39

4 Cl^(⊖) 40

4 Cl^(⊖) 41

4 Cl^(⊖) 42

Cl^(⊖) 43

Cl^(⊖) 44

3 Cl^(⊖) 45

3 Cl^(⊖) 46

4 Cl^(⊖) 47

3 Cl^(⊖)

TABLE 9

Example No. Q^(⊕) Y^(⊖) 48

Cl^(⊖) 49

2 Cl^(⊖) 50

3 Cl^(⊖) 51

3 Cl^(⊖) 52

3 Cl^(⊖) 53

3 Cl^(⊖)

TABLE 10

Example No. Q^(⊕) Y^(⊖) 54

Cl^(⊖) 55

2 Cl^(⊖) 56

3 Cl^(⊖) 57

3 Cl^(⊖) 58

3 Cl^(⊖) 59

4 Cl^(⊖)

Example No. Q^(⊕) Y^(⊖) 60

Cl^(⊖) 61

2 Cl^(⊖) 62

3 Cl^(⊖) 63

3 Cl^(⊖) 64

3 Cl^(⊖) 65

4 Cl^(⊖)

TABLE 2

Example No. Q^(⊕) Y^(⊖) 66

Cl^(⊖) 67

2 Cl^(⊖) 68

3 Cl^(⊖) 69

3 Cl^(⊖) 70

3 Cl^(⊖) 71

4 Cl^(⊖)

TABLE 13

Example No. Q^(⊕) Y^(⊖) 72

Cl^(⊖) 73

2 Cl^(⊖) 74

3 Cl^(⊖) 75

3 Cl^(⊖) 76

3 Cl^(⊖) 77

4 Cl^(⊖)

TABLE 14

Example No. Q^(⊕) Y^(⊖) 78

Cl^(⊖) 79

2 Cl^(⊖) 80

3 Cl^(⊖) 81

3 Cl^(⊖) 82

3 Cl^(⊖) 83

4 Cl^(⊖)

TABLE 15

Example No. Q^(⊕) Y^(⊖) 84

Cl^(⊖) 85

2 Cl^(⊖) 86

3 Cl^(⊖) 87

3 Cl^(⊖) 88

3 Cl^(⊖) 89

4 Cl^(⊖)

TABLE 16

Example No. Q^(⊕) Y^(⊖) 90

Cl^(⊖) 91

2 Cl^(⊖) 92

3 Cl^(⊖) 93

3 Cl^(⊖) 94

3 Cl^(⊖) 95

4 Cl^(⊖)

TABLE 17

Example No. Q^(⊕) Y^(⊖) 96

Cl^(⊖) 97

2 Cl^(⊖) 98

3 Cl^(⊖) 99

3 Cl^(⊖) 100

3 Cl^(⊖) 101

4 Cl^(⊖)

TABLE 18 Example No. 102

103

104


23. A pharmaceutical composition comprised of a compound in accordancewith claim 1 in combination with a pharmaceutically acceptable carrier.24. A pharmaceutical composition in accordance with claim 23 used in themanufacture of a medicament for the treatment of bacterial infections.25. A pharmaceutical composition in accordance with claim 23 furthercomprising a β-lactam antibiotic.
 26. A pharmaceutical composition inaccordance with claim 25 wherein the β-lactam is a carbapenemantibiotic.
 27. A composition according to claim 25 which furthercontains a serine β-lactamase inhibitor.
 28. A composition according toclaim 26 which further contains a DHP inhibitor.
 29. A method oftreating a bacterial infection comprising administering to a mammalianpatient in need of such treatment a metallo-β-lactamase inhibitorcompound as defined in claim 1 in combination with a pharmaceuticallyacceptable β-lactam antibiotic in an amount which is effective fortreating a bacterial infection.
 30. A method according to claim 29wherein the β-lactam is a carbapenem antibiotic.
 31. A method accordingto claim 30, which further contains a DHP inhibitor.
 32. A methodaccording to claim 31 wherein the DHP inhibitor is cilastatin.
 33. Amethod according to claim 29 which further contains a serine β-lactamaseinhibitor.